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Sample records for carcinoma cells down-regulation

  1. Down-regulation of vimentin expression inhibits carcinoma cell migration and adhesion.

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    McInroy, Lorna; Määttä, Arto

    2007-08-17

    Vimentin is a type III Intermediate filament protein that is expressed frequently in epithelial carcinomas correlating with invasiveness and poor prognosis. We have analysed migration and adhesion to collagenous matrix of a panel of carcinoma cell lines. In vitro invasiveness was highest in vimentin-positive SW480 colon cancer and MDA-MB-231 breast cancer cells and the role of vimentin in these cell lines was investigated by RNA interference. Down-regulation of vimentin expression resulted in impaired migration in both scratch-wound experiments and in invasion assays through cell culture inserts coated with collagen gel. Compromised migration was observed in both cell lines, whereas cell attachment assays revealed impaired adhesion to fibrillar collagen in MDA-MB-231 cells while the adhesion of vimentin-ablated SW480 cells, that express both vimentin and keratin intermediate filaments was not affected. In conclusion, ablation of vimentin expression inhibits migration and invasion of colon and breast cancer cell lines.

  2. Slug down-regulation by RNA interference inhibits invasion growth in human esophageal squamous cell carcinoma

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    Zhang Shaoyan

    2011-05-01

    Full Text Available Abstract Background Esophageal squamous cell carcinoma (ESCC is one of the most aggressive carcinomas of the gastrointestinal tract. We assessed the relevance of Slug in measuring the invasive potential of ESCC cells in vitro and in vivo in immunodeficient mice. Methods We utilized RNA interference to knockdown Slug gene expression, and effects on survival and invasive carcinoma were evaluated using a Boyden chamber transwell assay in vitro. We evaluated the effect of Slug siRNA-transfection and Slug cDNA-transfection on E-cadherin and Bcl-2 expression in ESCC cells. A pseudometastatic model of ESCC in immunodeficient mice was used to assess the effects of Slug siRNA transfection on tumor metastasis development. Results The EC109 cell line was transfected with Slug-siRNA to knockdown Slug expression. The TE13 cell line was transfected with Slug-cDNA to increase Slug expression. EC109 and TE13 cell lines were tested for the expression of apoptosis-related genes bcl-2 and metastasis-related gene E-cadherin identified previously as Slug targets. Bcl-2 expression was increased and E-cadherin was decreased in Slug siRNA-transfected EC109 cells. Bcl-2 expression was increased and E-cadherin was decreased in Slug cDNA-transfected TE13 cells. Invasion of Slug siRNA-transfected EC109 cells was reduced and apoptosis was increased whereas invasion was greater in Slug cDNA-transfected cells. Animals injected with Slug siRNA-transfected EC109 cells exhihited fewer seeded nodes and demonstrated more apoptosis. Conclusions Slug down-regulation promotes cell apoptosis and decreases invasion capability in vitro and in vivo. Slug inhibition may represent a novel strategy for treatment of metastatic ESCC.

  3. Andrographolide could inhibit human colorectal carcinoma Lovo cells migration and invasion via down-regulation of MMP-7 expression.

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    Shi, Ming-Der; Lin, Hui-Hsuan; Chiang, Tai-An; Tsai, Li-Yu; Tsai, Shu-Mei; Lee, Yi-Chieh; Chen, Jing-Hsien

    2009-08-14

    Andrographolide (Andro), a diterpenoid lactone isolated from a traditional herbal medicine Andrographis paniculata, is known to possess multiple pharmacological activities. In our previous study, Andro had been shown to have potent anti-cancer activity against human colorectal carcinoma Lovo cells by inhibiting cell-cycle progression. To further investigate the mechanism for the anti-cancer properties of Andro, it was used to examine the effect on migration and invasion of Lovo cells. The results of wound-healing assay and in vitro transwell assay revealed that Andro inhibited dose-dependently the migration and invasion of Lovo cells under non-cytotoxic concentrations. Using zymographic assay and RT-PCR, the results revealed that Andro diminished the activity and the mRNA and protein levels of MMP-7, but not MMP-2 or MMP-9. The down-regulation of MMP-7 appeared to be via the inactivation of activator protein-1 (AP-1) since the treatment with Andro suppressed the nuclear protein level of AP-1, which was accompanied by a decrease in DNA-binding level of the factor. Taken together, these results indicated that Andro reduces the MMP-7-mediated cellular events in Lovo cells, and provided a new mechanism for its anti-cancer activity.

  4. Down-regulation of cytoplasmic PLZF correlates with high tumor grade and tumor aggression in non-small cell lung carcinoma.

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    Xiao, Guang-Qian; Li, Faqian; Findeis-Hosey, Jennifer; Hyrien, Ollivier; Unger, Pamela D; Xiao, Lu; Dunne, Richard; Kim, Eric S; Yang, Qi; McMahon, Loralee; Burstein, David E

    2015-11-01

    There are currently no effective prognostic biomarkers for lung cancer. Promyelocytic leukemia zinc finger (PLZF), a transcriptional repressor, has a role in cell cycle progression and tumorigenicity in various cancers. The expression and value of PLZF in lung carcinoma, particularly in the subclass of non-small cell lung carcinoma (NSCLC), has not been studied. Our aim was to study the immunohistochemical expression of PLZF in lung adenocarcinoma and squamous cell carcinoma and correlate the alteration of PLZF expression with tumor differentiation, lymph node metastasis, tumor stage, and overall survival. A total of 296 NSCLCs being mounted on tissue microarray (181 adenocarcinomas and 91 squamous cell carcinomas) were investigated. Moderate to strong expression of PLZF was found in the cytoplasm of all the nonneoplastic respiratory epithelium and most (89.9%) well-differentiated adenocarcinoma. The proportions of moderately differentiated, poorly differentiated adenocarcinoma, and paired lymph node adenocarcinoma metastases that demonstrated negative or only weak PLZF reactivity were 75.6%, 97.2%, and 89.9%, respectively. The expression of PLZF in squamous cell carcinoma was mostly weak or absent and significantly lower than that in adenocarcinoma of the same grade (P carcinoma and adenocarcinoma (P < .0001). Down-regulation of PLZF also correlated with higher tumor stage and shorter overall survival (P < .05). These results support a prognostic value for loss of cytoplasmic PLZF expression in the stratification of NSCLC and a possible role of cytoplasmic shift and down-regulation of PLZF in the pathogenesis of NSCLC.

  5. Long Non-Coding RNA HOTAIR Promotes Cell Migration and Invasion via Down-Regulation of RNA Binding Motif Protein 38 in Hepatocellular Carcinoma Cells

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    Chaofeng Ding

    2014-03-01

    Full Text Available Long non-coding RNA HOTAIR exerts regulatory functions in various biological processes in cancer cells, such as proliferation, apoptosis, mobility, and invasion. We previously found that HOX transcript antisense RNA (HOTAIR is a negative prognostic factor and exhibits oncogenic activity in hepatocellular carcinoma (HCC. In this study, we aimed to investigate the role and molecular mechanism of HOTAIR in promoting HCC cell migration and invasion. Firstly, we profiled its gene expression pattern by microarray analysis of HOTAIR loss in Bel-7402 HCC cell line. The results showed that 129 genes were significantly down-regulated, while 167 genes were significantly up-regulated (fold change >2, p < 0.05. Bioinformatics analysis indicated that RNA binding proteins were involved in this biological process. HOTAIR suppression using RNAi strategy with HepG2 and Bel-7402 cells increased the mRNA and protein expression levels of RNA binding motif protein 38 (RBM38. Moreover, the expression levels of RBM38 in HCC specimens were significantly lower than paired adjacent noncancerous tissues. In addition, knockdown of HOTAIR resulted in a decrease of cell migration and invasion, which could be specifically rescued by down-regulation of RBM38. Taken together, HOTAIR could promote migration and invasion of HCC cells by inhibiting RBM38, which indicated critical roles of HOTAIR and RBM38 in HCC progression.

  6. Expression of DIAPH1 is up-regulated in colorectal cancer and its down-regulation strongly reduces the metastatic capacity of colon carcinoma cells.

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    Lin, Yuan-Na; Izbicki, Jakob R; König, Alexandra; Habermann, Jens K; Blechner, Christine; Lange, Tobias; Schumacher, Udo; Windhorst, Sabine

    2014-04-01

    In most cases, metastatic colorectal cancer is not curable, thus new approaches are necessary to identify novel targets for colorectal cancer therapy. Actin-binding-proteins (ABPs) directly regulate motility of metastasising tumor cells, and for cortactin an association with colon cancer metastasis has been already shown. However, as its depletion only incompletely inhibits metastasis, additional, more suitable cellular targets have to be identified. Here we analyzed expression of the ABPs, DIAPH1, VASP, N-WASP, and fascin in comparison with cortactin and found that, besides cortactin, DIAPH1 was expressed with the highest frequency (63%) in colorectal cancer. As well as cortactin, DIAPH1 was not detectable in normal colon tissue and expression of both proteins was positively correlated with metastasis of colorectal cancer. To analyse the mechanistic role of DIAPH1 for metastasis of colon carcinoma cells in comparison with cortactin, expression of the proteins was stably down-regulated in the human colon carcinoma cell lines HT-29, HROC-24 and HCT-116. Analysis of metastasis of colon carcinoma cells in SCID mice revealed that depletion of DIAPH1 reduced metastasis 60-fold and depletion of cortactin 16-fold as compared with control cells. Most likely the stronger effect of DIAPH1 depletion on colon cancer metastasis is due to the fact that in vitro knock down of DIAPH1 impaired all steps of metastasis; adhesion, invasion and migration while down-regulation of cortactin only reduced adhesion and invasion. This very strong reducing effect of DIAPH1 depletion on colon carcinoma cell metastasis makes the protein a promising therapeutic target for individualized colorectal cancer therapy.

  7. mad—overexpression down regulates the malignant growth and p53 mediated apoptosis in human hepatocellular carcinoma BEL—7404 cells

    Institute of Scientific and Technical Information of China (English)

    ZHANHUA; YONGHUAXU

    1999-01-01

    Mad protein has been shown as an antagonist of cMyc protein in some cell lines.The effect of Mad protein to the malignant phenotype of human hepatoma BEL-7404 cell line was investigated experimentally.An eukarryotic vector pCDNA Ⅲ containing full ORF fragment of mad cDNA was transfected into targeted cells.Under G418 selection,stable Mad-overexpressed cells were cloned.Studies on the effect of Mad over-expression in cell proliferation and cell cycle revealed that cell morphology of the Mad-overexpressed BEL-7404-M1 cells was significantly different from the parent and control vector transfected cells.DNA synthesis,cell proliferation and anchorage-independent growth in soft-agar of the madtransfected cells were partially inhibited in comparison to control cells.Flos cytometry analysis indicated that mad over-expression might block more transfectant cells at G0/G1 phase,resulting in the retardation of cell proliferation.RT-PCR detected a marked inhibition of the expression of cdc25A,an important regulator gene of G0/G1 to S phase in cell cycle.It was also found that Mad protein overexpression could greatly suppress p53-mediated apoptosis in BEL-74040M1 cells in the absence of serume.Thus,Mad proteins may function as a negative regulator antagonizing c-Myc activity in the control of cell growth and apoptosis in human hepatocellular carcinoma BEL-7404 cells.

  8. Down-regulation of EBV-LMP1 radio-sensitizes nasal pharyngeal carcinoma cells via NF-κB regulated ATM expression.

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    Xiaoqian Ma

    Full Text Available BACKGROUND: The latent membrane protein 1 (LMP1 encoded by EBV is expressed in the majority of EBV-associated human malignancies and has been suggested to be one of the major oncogenic factors in EBV-mediated carcinogenesis. In previous studies we experimentally demonstrated that down-regulation of LMP1 expression by DNAzymes could increase radiosensitivity both in cells and in a xenograft NPC model in mice. RESULTS: In this study we explored the molecular mechanisms underlying the radiosensitization caused by the down-regulation of LMP1 in nasopharyngeal carcinoma. It was confirmed that LMP1 could up-regulate ATM expression in NPCs. Bioinformatic analysis of the ATM ptomoter region revealed three tentative binding sites for NF-κB. By using a specific inhibitor of NF-κB signaling and the dominant negative mutant of IkappaB, it was shown that the ATM expression in CNE1-LMP1 cells could be efficiently suppressed. Inhibition of LMP1 expression by the DNAzyme led to attenuation of the NF-κB DNA binding activity. We further showed that the silence of ATM expression by ATM-targeted siRNA could enhance the radiosensitivity in LMP1 positive NPC cells. CONCLUSIONS: Together, our results indicate that ATM expression can be regulated by LMP1 via the NF-κB pathways through direct promoter binding, which resulted in the change of radiosensitivity in NPCs.

  9. Down-regulation of microRNAs controlling tumourigenic factors in follicular thyroid carcinoma

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    Rossing, Maria; Helweg-Larsen, Rehannah Borup; Henao Giraldo, Ricardo

    2012-01-01

    ) and follicular carcinoma (FC). Comparison of carcinoma and adenoma with normal thyroid revealed 150 and 107 differentially expressed miRNAs. Most miRNAs were down-regulated and especially miR-199b-5p and miR-144 which were essentially lost in the carcinomas. Integration of the changed miRNAs with differentially...... expressed mRNAs demonstrated an enrichment of seed-sites among up-regulated transcripts encoding proteins implicated in thyroid tumourigenesis. This was substantiated by the demonstration that pre-miR-199b reduced proliferation when added to cultured follicular thyroid carcinoma cells. The down-regulated mi......RNAs in FC exhibited a substantial similarity with down-regulated miRNAs in anaplastic carcinoma and by gene set enrichment analysis, we observed a significant identity between target mRNAs in FC and transcripts up-regulated in anaplastic carcinoma. To examine the diagnostic potential of miRNA expression...

  10. Down-regulation of UDP-glucose dehydrogenase affects glycosaminoglycans synthesis and motility in HCT-8 colorectal carcinoma cells

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    Wang, Tsung-Pao; Pan, Yun-Ru; Fu, Chien-Yu; Chang, Hwan-You, E-mail: hychang@life.nthu.edu.tw

    2010-10-15

    UDP-glucose dehydrogenase (UGDH) catalyzes oxidation of UDP-glucose to yield UDP-glucuronic acid, a precursor of hyaluronic acid (HA) and other glycosaminoglycans (GAGs) in extracellular matrix. Although association of extracellular matrix with cell proliferation and migration has been well documented, the importance of UGDH in these behaviors is not clear. Using UGDH-specific small interference RNA to treat HCT-8 colorectal carcinoma cells, a decrease in both mRNA and protein levels of UGDH, as well as the cellular UDP-glucuronic acid and GAG production was observed. Treatment of HCT-8 cells with either UGDH-specific siRNA or HA synthesis inhibitor 4-methylumbelliferone effectively delayed cell aggregation into multicellular spheroids and impaired cell motility in both three-dimensional collagen gel and transwell migration assays. The reduction in cell aggregation and migration rates could be restored by addition of exogenous HA. These results indicate that UGDH can regulate cell motility through the production of GAG. The enzyme may be a potential target for therapeutic intervention of colorectal cancers.

  11. Tivantinib (ARQ-197) exhibits anti-tumor activity with down-regulation of FAK in oral squamous cell carcinoma

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    Xi, Wei-Hong [Department of Oral and Maxillofacial Surgery, First Affiliated Hospital, Nanchang University, Nanchang 330006 (China); Yang, Li-Yun [Department of Blood Transfusion, First Affiliated Hospital, Nanchang University, Nanchang 330006 (China); Cao, Zhong-Yi, E-mail: m18070383032@163.com [Department of Oral and Maxillofacial Surgery, First Affiliated Hospital, Nanchang University, Nanchang 330006 (China); Qian, Yong, E-mail: yfykqkqy@163.com [Department of Oral and Maxillofacial Surgery, First Affiliated Hospital, Nanchang University, Nanchang 330006 (China)

    2015-02-20

    Oral squamous cell carcinoma (OSCC) is one of the most common cancers worldwide and the 5 years survival rate of the patients is about 60% in the USA, due to acquired chemotherapeutic resistance and metastasis of the disease. In this study, we found that tivantinib, a selective MET inhibitor, suppresses OCSS cell proliferation and colony formation, however, anti-tumor activities induced by tivantinib are independent of the inhibition of MET signaling pathway. In addition, tivantinib cause G2/M cell cycle arrest and caspases-dependent apoptosis in OSCC cell lines. We also found that tivantinib dose-dependently suppressed the activation and expression of FAK. In all, these data suggested that tivantinib may be developed as a chemotherapeutic agent to effectively treat certain cancers including OSCC. - Highlights: • Tivantinib suppresses OSCC cell growth independent of the inhibition of HGF/MET signaling pathway. • Tivantinib blocks cell cycle and induces caspases-mediated apoptosis. • Tivantinib elicits its anti-tumor activity with the inhibition of FAK signaling pathway.

  12. Prokineticin 2/Bv8 is expressed in Kupffer cells in liver and is down regulated in human hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    AIM: To study the implication of prokineticin 1 (PK1/EG VEGF) and prokineticin 2 (PK2/Bv8) in hepatocellular carcinoma angiogenesis. METHODS: The gene induction of PK1/EG-VEGF and PK2/Bv8 was investigated in 10 normal, 28 fibrotic and 28 tumoral livers by using real time PCR. Their expression was compared to the expression of VEGF (an angiogenesis marker), vWF (an endothelial cell marker) and to CD68 (a monocyte/macrophage marker). Furthermore, the mRNA levels of PK1/EG-VEGF, PK2/Bv8, prokineticin receptor 1 and 2 were evaluated by real time PCR in isolated liver cell populations. Finally, PK2/Bv8protein was detected in normal liver paraffin sections and in isolated liver cells by immunohistochemistry and immunocytochemistry.RESULTS:PK2/Bv8 mRNA but not PK1/EG-VEGF was expressed in all types of normal liver samples examined. In the context of liver tumor development, we reported that PK2/Bv8 correlates only with CD68 and showed a significant decrease in expression as the pathology evolves towards cancer. Whereas, VEGF and vWF mRNA were significantly upregulated in both fibrosis and HCC, as expected. In addition, out of all isolated liver cells examined, only Kupffer cells (liver resident macrophages) express significant levels of PK2/Bv8 and its receptors, prokineticin receptor 1 and 2.CONCLUSION:In normal liver PK2/Bv8 and its receptors were specifically expressed by Kupffer cells. PK2/Bv8 expression decreased as the liver evolves towards cancer and did not correlate with HCC angiogenesis.

  13. Proteomic Identification of LASP-1 Down-regulation After RNAi Urokinase Silencing in Human Hepatocellular Carcinoma Cells

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    Alessandro Salvi

    2009-02-01

    Full Text Available In human hepatocellular carcinoma (HCC, the high expression of urokinase-type plasminogen activator (uPA is an unfavorable prognostic factor and a therapeutic target. To identify the downstream effects of uPA silencing by RNA interference, we studied proteome modifications of uPA-inhibited SKHep1C3 cells, an HCC-derived cell line. The study with two-dimensional difference gel electrophoresis and matrix-assisted laser desorption/ionization-time of flight-mass spectrometry showed Lim and SH3 protein 1 (LASP-1, cytokeratin 1 (CK-1, cytokeratin 10 (CK-10, and heterogeneous nuclear ribonucleoprotein H1 down-modulation after uPA inhibition. LASP-1, CK-1, and CK-10 are involved in cytoskeleton dynamics as heterogeneous nuclear ribonucleoprotein H1 takes part in the mRNA processing and stability. We first confirmed the proteomic data by Western blot and immunoflorescence and then explored the link between uPA and LASP-1. The ectopic expression of uPA and LASP-1 supported the proteomic results and showed that uPA up-regulation increased LASP-1 expression and that both were implicated in SKHep1C3 motility. siRNA LASP-1 inhibition showed that LASP-1 was involved in actin microfilaments organization of SKHep1C3 cells. The disruption of the actin microfilaments after LASP-1 depletion increased uPA secretion and SKHep1C3 motility. Our results would suggest the hypothesis that uPA and LASP-1 expression may be coordinated in HCC-derived cells. In summary, the proteomic identification of a set of uPA downstream proteins provides new insight into the function of uPA in HCC cells.

  14. Solamargine inhibits migration and invasion of human hepatocellular carcinoma cells through down-regulation of matrix metalloproteinases 2 and 9 expression and activity.

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    Sani, Iman Karimi; Marashi, Seyed Hassan; Kalalinia, Fatemeh

    2015-08-01

    Solamargine is a steroidal alkaloid glycoside isolated from Solanum nigrum. The aim of this study was to investigate the effects of solamargine on tumor migration and invasion in aggressive human hepatocellular carcinoma cells. The MTT assay was used to assess the effects of solamargine on the viability of HepG2 cells. Migration and invasion ability of HepG2 cells under solamargine treatment were examined by a wound healing migration assay and Boyden chamber assay, respectively. Western blotting assays were used to detect the expression of MMP-2 and MMP-9 proteins and MMP-2 and MMP-9 activity were analyzed by gelatin zymography assay. Solamargine reduced HepG2 cell viability in a concentration-dependent manner. At 7.5μM solamargine decreased cell viability by less than 20% in HepG2 cells. A wound healing migration assay and Boyden chamber invasion assay showed that solamargine significantly inhibited in vitro migration and invasion of HepG2 cells. At the highest dose, solamargine decreased cell migration and invasion by more than 70% and 72% in HepG2 cells, respectively. Western blotting and gelatin zymography results showed that solamargine reduced expression and function of MMP-2 and MMP-9 proteins. In conclusion, the results showed that solamargine significantly inhibits migration and invasion of HepG2 cells by down-regulating MMP-2 and MMP-9 expression and activity.

  15. Protective Effects of Total Glucosides of Paeony on N-nitrosodiethylamine-induced Hepatocellular Carcinoma in Rats via Down-regulation of Regulatory B Cells.

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    Song, S S; Yuan, P F; Li, P P; Wu, H X; Ni, W J; Lu, J T; Wei, W

    2015-01-01

    Total glucoside of paeony (TGP), extracted from the root of Paeonia Lactiflora, has been known to show anti-inflammatory, anti-oxidative, hepato-protective and immuno-regulatory activities. The aim of this present study was to determine the anti-tumor effect of TGP against N-nitrosodiethylamine (DEN)-induced hepatocellular carcinoma (HCC) in rats, and to find the related mechanisms. Rat HCC model was established by intragastrically administrating with DEN (8 mg/kg). We found the number of tumor nodules and the index of liver and spleen were increased in the model group compared with the normal group, and was significantly decreased by TGP. Additionally, TGP obviously improved the hepatic pathological lesions induced by DEN, and decreased the elevated levels of serum alanine aminotransferase (ALT), glutamic oxalacetic transaminase (AST), alkaline phosphatase (ALP) and alpha fetoprotein (AFP) by DEN. Moreover, TGP decreased the level of B cell-activating factor (BAFF) and the proportion of IL-10-producing regulatory B cells (Bregs), and the decrease of BAFF by TGP is positively correlated to the decrease of IL-10-producing Bregs by TGP. These results suggest that TGP had a good therapeutic action on DEN-induced HCC rats, which might be due to its down-regulation of Bregs through reducing the level of BAFF.

  16. Down regulation of E-Cadherin (ECAD - a predictor for occult metastatic disease in sentinel node biopsy of early squamous cell carcinomas of the oral cavity and oropharynx

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    Studer Gabriela

    2011-06-01

    Full Text Available Abstract Background Prognostic factors in predicting occult lymph node metastasis in patients with head and neck squamous-cell carcinoma (HNSCC are necessary to improve the results of the sentinel lymph node procedure in this tumour type. The E-Cadherin glycoprotein is an intercellular adhesion molecule in epithelial cells, which plays an important role in establishing and maintaining intercellular connections. Objectives To determine the value of the molecular marker E-Cadherin in predicting regional metastatic disease. Methods E-Cadherin expression in tumour tissue of 120 patients with HNSCC of the oral cavity and oropharynx were evaluated using the tissue microarray technique. 110 tumours were located in the oral cavity (91.7%; mostly tongue, 10 tumours in the oropharynx (8.3%. Intensity of E-Cadherin expression was quantified by the Intensity Reactivity Score (IRS. These results were correlated with the lymph node status of biopsied sentinel lymph nodes. Univariate and multivariate analysis was used to determine statistical significance. Results pT-stage, gender, tumour side and location did not correlate with lymph node metastasis. Differentiation grade (p = 0.018 and down regulation of E-Cadherin expression significantly correlate with positive lymph node status (p = 0.005 in univariate and multivariate analysis. Conclusion These data suggest that loss of E-cadherin expression is associated with increased lymhogeneous metastasis of HNSCC. E-cadherin immunohistochemistry may be used as a predictor for lymph node metastasis in squamous cell carcinoma of the oral cavity and oropharynx. Level of evidence: 2b

  17. Curcumin Promoted the Apoptosis of Cisplain-resistant Human Lung Carcinoma Cells A549/DDP through Down-regulating miR-186*

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    Jian ZHANG

    2010-04-01

    Full Text Available Background and objective Curcumin, a natural compound, is derived from the rthizom of Curcuma longa. In vitro and in vivo preclinical studies have shown its anti-inflammatory, antioxidant, anticancer activities and so on. miR-186*, which was found by microarray technology, was highly expressed in lung carcinoma cells A549/DDP. The aim of this study is to illustrate whether Curcumin could promote the apoptosis of A549/DDP cells through regulating the expression of miR-186*. Methods An oligonucleotide microarray chip was used to profile microRNA (miRNA expressions in A549/DDP cells treated with and without Curcumin. The significantly differentially expressed miRNA, which was selected from microarray chip, validated by quantitative real-time PCR. Ultimately, the remarkably expressed miRNA modulated the apoptosis assaying by flow cytometry expriments and the survival rate was measured by MTT method. Results The microarray chip results demonstrated: Curcumin altered the expression level of miRNAs compared with untreated control in A549/DDP cell line, miR-186* was significantly down-regulated after Curcumin treatment, which confirmed by quantitative real-time PCR. Downregulation of miR-186* expression by curcumin elevated the apoptosis, and the survival rate of A549/DDP cells decreased; but up-regulation of miR-186* expression by transfection its mimics restrained the apoptosis, the survival rate of A549/DDP cells increased, which were assayed by flow cytometry expriments and MTT method. Conclusion Modulation of miRNAs expression may be an important mechanism underlying the biological roles of Curcumin.

  18. Down-regulated miR-9 and miR-433 in human gastric carcinoma

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    Nie Na

    2009-06-01

    Full Text Available Abstract Background MircoRNAs(miRNAs are short, endogenously non-coding RNAs. The abnormal expression of miRNAs may be valuable for the diagnosis and treatment of tumors. Methods To screening the special miRNAs in gastric carcinoma, expression level of miRNAs in gastric carcinoma and normal gaster samples were detected by miRNA gene chip. Then, the expressions of miR-9 and miR-433 in gastric carcinoma tissue and SGC7901 cell line were validated by qRT-PCR. GRB2 and RAB34, targets of miR-433 and miR-9 respectively, were detected by Western blot. Results We found 19 miRNAs and 7 miRNAs were down-regulated and up-regulated respectively. Compared with normal gaster samples, our data showed that miR-9 and miR-433 were down-regulated in gastric carcinoma. Meanwhile, we also found that miR-433 and miR-9 regulated the expression levels of GRB2 and RAB34 respectively. Conclusion Our data show miR-9 and miR-433 was down-regulated in gastric carcinoma. The targets of miR-433 and miR-9 were tumor-associated proteins GRB2 and RAB34 respectively. This result provided the related information of miRNAs in gastric carcinoma.

  19. MicroRNA-21 promotes cell proliferation and down-regulates the expression of programmed cell death 4 (PDCD4) in HeLa cervical carcinoma cells

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    Yao, Qing; Xu, Hui; Zhang, Qian-Qian; Zhou, Hui [Key Laboratory of Gene Engineering of the Ministry of Education, State Key Laboratory for Biocontrol, Sun Yat-Sen University, Guangzhou 510275 (China); Qu, Liang-Hu, E-mail: lssqlh@mail.sysu.edu.cn [Key Laboratory of Gene Engineering of the Ministry of Education, State Key Laboratory for Biocontrol, Sun Yat-Sen University, Guangzhou 510275 (China)

    2009-10-23

    MicroRNAs are involved in cancer-related processes. The microRNA-21(miR-21) has been identified as the only miRNA over-expressed in a wide variety of cancers, including cervical cancer. However, the function of miR-21 is unknown in cervical carcinomas. In this study, we found that the inhibition of miR-21 in HeLa cervical cancer cells caused profound suppression of cell proliferation, and up-regulated the expression of the tumor suppressor gene PDCD4. We also provide direct evidence that PDCD4-3'UTR is a functional target of miR-21 and that the 18 bp putative target site can function as the sole regulatory element in HeLa cells. These results suggest that miR-21 may play an oncogenic role in the cellular processes of cervical cancer and may serve as a target for effective therapies.

  20. Afatinib down-regulates MCL-1 expression through the PERK-eIF2α-ATF4 axis and leads to apoptosis in head and neck squamous cell carcinoma.

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    Liu, Xianfang; Lv, Zhenghua; Zou, Jidong; Liu, Xiuxiu; Ma, Juke; Wang, Jinhua; Sa, Na; Jing, Peihang; Xu, Wei

    2016-01-01

    Afatinib is the second generation of irreversible inhibitor of EGFR, HER2 and HER4, which has shown encouraging phase II and III clinical outcomes in the treatment of head and neck squamous cell carcinoma (HNSCC). However, the molecular mechanism of afatinib-induced apoptosis in HNSCC is poorly understood. In the present investigation, we discovered that down-regulation of MCL-1, an anti-apoptotic member of BCL-2 family, was responsible for afatinib-triggered apoptosis. And the inactivation of AKT-mTOR signaling caused by afatinib lead to translational inhibition of MCL-1 expression. As a crucial branch of ER stress, PERK-eIF2α-ATF4 axis was also stimulated in HNSCC cells after afatinib incubation. Silencing either eIF2α or ATF4 by siRNA transfection relieved afatinib-caused suppression of AKT-mTOR activity, attenuating MCL-1 down-regulation as well as subsequent apoptosis. Collectively, the results show that afatinib hampers AKT-mTOR activation by stimulating PERK-eIF2α-ATF4 signaling pathway, giving rise to MCL-1 down-regulation mediated apoptosis in HNSCC cells. Therefore, our findings reveal the elaborate molecular network of afatinib-induced apoptosis in HNSCC, which would provide substantial theoretical underpinnings for afatinib clinical application and highlight its promising prospect in HNSCC treatment.

  1. Physapubescin selectively induces apoptosis in VHL-null renal cell carcinoma cells through down-regulation of HIF-2α and inhibits tumor growth

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    Chen, Lixia; Xia, Guiyang; Qiu, Feng; Wu, Chunli; Denmon, Andria P.; Zi, Xiaolin

    2016-01-01

    We have purified physapubescin, a predominant steroidal lactone, from medicinal plant Physalis pubescens L., commonly named as “hairy groundcherry” in English and “Deng-Long-Cao” in Chinese. Von Hippel-Lindau (VHL)-null 786-O, RCC4 and A498 Renal Cell Carcinoma (RCC) cell lines expressing high levels of Hypoxia Inducible Factor (HIF)-2α are more sensitive to physapubescin-mediated apoptosis and growth inhibitory effect than VHL wild-type Caki-2 and ACHN RCC cell lines. Restoration of VHL in RCC4 cells attenuated the growth inhibitory effect of physapubescin. Physapubescin decreases the expression of HIF-2α and increases the expression of CCAAT/enhancer-binding protein homologus protein (CHOP), which leads to up-regulation of death receptor 5 (DR5), activation of caspase-8 and -3, cleavage of poly (ADP-Ribose) polymerase (PARP) and apoptosis. Under hypoxia conditions, the apoptotic and growth inhibitory effects of physapubescin are further enhanced. Additionally, physapubescin synergizes with TNF-related apoptosis-inducing ligand (TRAIL) for markedly enhanced induction of apoptosis in VHL-null 786-O cells but not in VHL wild-type Caki-2 cells. Physapubescin significantly inhibited in vivo angiogenesis in the 786-O xenograft. Physapubescin as a novel agent for elimination of VHL-null RCC cells via apoptosis is warranted for further investigation. PMID:27581364

  2. RNA interference targeting CD147 inhibits the proliferation, invasiveness, and metastatic activity of thyroid carcinoma cells by down-regulating glycolysis.

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    Huang, Peng; Chang, Shi; Jiang, Xiaolin; Su, Juan; Dong, Chao; Liu, Xu; Yuan, Zhengtai; Zhang, Zhipeng; Liao, Huijun

    2015-01-01

    A high rate of glycolytic flux, even in the presence of oxygen, is a key metabolic hallmark of cancer cells. Lactate, the end product of glycolysis, decreases the extracellular pH and contributes to the proliferation, invasiveness and metastasis of tumor cells. CD147 play a crucial role in tumorigenicity, invasion and metastasis; and CD147 also interacts strongly and specifically with monocarboxylate transporter1 (MCT1) that mediates the transport of lactate. The objective of this study was to determine whether CD147 is involved, via its association with MCT1 to transport lactate, in glycolysis, contributing to the progression of thyroid carcinoma. The expression levels of CD147 in surgical specimens of normal thyroid, nodular goiter (NG), well-differentiated thyroid carcinoma (WDTC), and undifferentiated thyroid carcinoma (UDTC) were determined using immunohistochemical techniques. The effects of CD147 silencing on cell proliferation, invasiveness, metastasis, co-localization with MCT1, glycolysis rate and extracellular pH of thyroid cancer cells (WRO and FRO cell lines) were measured after CD147 was knocked-down using siRNA targeting CD147. Immunohistochemical analysis of thyroid carcinoma (TC) tissues revealed significant increases in signal for CD147 compared with normal tissue or NG, while UDTC expressed remarkably higher levels of CD147 compared with WDTC. Furthermore, silencing of CD147 in TC cells clearly abrogated the expression of MCT1 and its co-localization with CD147 and dramatically decreased both the glycolysis rate and extracellular pH. Thus, cell proliferation, invasiveness, and metastasis were all significantly decreased by siRNA. These results demonstrate in vitro that the expression of CD147 correlates with the degree of dedifferentiation of thyroid cancer, and show that CD147 interacts with MCT1 to regulate tumor cell glycolysis, resulting in the progression of thyroid carcinoma.

  3. Up-Regulation of PAI-1 and Down-Regulation of uPA Are Involved in Suppression of Invasiveness and Motility of Hepatocellular Carcinoma Cells by a Natural Compound Berberine.

    Science.gov (United States)

    Wang, Xuanbin; Wang, Ning; Li, Hongliang; Liu, Ming; Cao, Fengjun; Yu, Xianjun; Zhang, Jingxuan; Tan, Yan; Xiang, Longchao; Feng, Yibin

    2016-04-16

    Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death and its prognosis remains poor due to the high risk of tumor recurrence and metastasis. Berberine (BBR) is a natural compound derived from some medicinal plants, and accumulating evidence has shown its potent anti-tumor activity with diverse action on tumor cells, including inducing cancer cell death and blocking cell cycle and migration. Molecular targets of berberine involved in its inhibitory effect on the invasiveness remains not yet clear. In this study, we identified that berberine exhibits a potent inhibition on the invasion and migration of HCC cells. This was accompanied by a dose-dependent down-regulation of expression of Cyclooxygenase-2 (COX-2), nuclear factor kappa B (NF-κB), urokinase-type plasminogen activator (uPA) and matrix metalloproteinase (MMP)-9 in berberine-treated HCC cells. Furthermore, berberine inactivated p38 and Erk1/2 signaling pathway in HCC cells. Primarily, this may be attributed to the up-regulation of plasminogen activator inhibitor-1 (PAI-1), a tumor suppressor that can antagonize uPA receptor and down-regulation of uPA. Blockade of uPA receptor-associated pathways leads to reduced invasiveness and motility of berberine-treated HCC cells. In conclusion, our findings identified for the first time that inactivation of uPA receptor by up-regulation of PAI-1 and down-regulation of uPA is involved in the inhibitory effect of berberine on HCC cell invasion and migration.

  4. Silibinin Suppresses Growth of Human Colorectal Carcinoma SW480 Cells in Culture and Xenograft through Down-regulation of β-Catenin-Dependent Signaling

    Directory of Open Access Journals (Sweden)

    Manjinder Kaur

    2010-05-01

    Full Text Available Mutations in APC/β-catenin resulting in an aberrant activation of Wnt/β-catenin pathway are common in colorectal cancer (CRC, suggesting that targeting the β-catenin pathway with chemopreventive/anticancer agents could be a potential translational approach to control CRC. Using human CRC cell lines harboring mutant (SW480 versus wildtype (HCT116 APC gene and alteration in β-catenin pathway, herein we performed both in vitro and in vivo studies to examine for the first time whether silibinin targets β-catenin pathway in its efficacy against CRC. Silibinin treatment inhibited cell growth, induced cell death, and decreased nuclear and cytoplasmic levels of β-catenin in SW480 but not in HCT116 cells, suggesting its selective effect on the β-catenin pathway and associated biologic responses. Other studies, therefore, were performed only in SW480 cells where silibinin significantly decreased β-catenin-dependent T-cell factor-4 (TCF-4 transcriptional activity and protein expression of β-catenin target genes such as c-Myc and cyclin D1. Silibinin also decreased cyclin-dependent kinase 8 (CDK8, a CRC oncoprotein that positively regulates β-catenin activity, and cyclin C expression. In a SW480 tumor xenograft study, 100- and 200-mg/kg doses of silibinin feeding for 6 weeks inhibited tumor growth by 26% to 46% (P < .001. Analyses of xenografts showed that similar to cell culture findings, silibinin decreases proliferation and expression of β-catenin, cyclin D1, c-Myc, and CDK8 but induces apoptosis in vivo. Together, these findings suggest that silibinin inhibits the growth of SW480 tumors carrying the mutant APC gene by down-regulating CDK8 and β-catenin signaling and, therefore, could be an effective agent against CRC.

  5. Induction of apoptosis in renal cell carcinoma by reactive oxygen species: involvement of extracellular signal-regulated kinase 1/2, p38delta/gamma, cyclooxygenase-2 down-regulation, and translocation of apoptosis-inducing factor.

    LENUS (Irish Health Repository)

    Ambrose, Monica

    2012-02-03

    Renal cell carcinoma (RCC) is the most common malignancy of the kidney. Unfortunately, RCCs are highly refractory to conventional chemotherapy, radiation therapy, and even immunotherapy. Thus, novel therapeutic targets need to be sought for the successful treatment of RCCs. We now report that 6-anilino-5,8-quinolinequinone (LY83583), an inhibitor of cyclic GMP production, induced growth arrest and apoptosis of the RCC cell line 786-0. It did not prove deleterious to normal renal epithelial cells, an important aspect of chemotherapy. To address the cellular mechanism(s), we used both genetic and pharmacological approaches. LY83583 induced a time- and dose-dependent increase in RCC apoptosis through dephosphorylation of mitogen-activated protein kinase kinase 1\\/2 and its downstream extracellular signal-regulated kinases (ERK) 1 and -2. In addition, we observed a decrease in Elk-1 phosphorylation and cyclooxygenase-2 (COX-2) down-regulation. We were surprised that we failed to observe an increase in either c-Jun NH(2)-terminal kinase or p38alpha and -beta mitogen-activated protein kinase activation. In contradiction, reintroduction of p38delta by stable transfection or overexpression of p38gamma dominant negative abrogated the apoptotic effect. Cell death was associated with a decrease and increase in Bcl-x(L) and Bax expression, respectively, as well as release of cytochrome c and translocation of apoptosis-inducing factor. These events were associated with an increase in reactive oxygen species formation. The antioxidant N-acetyl l-cysteine, however, opposed LY83583-mediated mitochondrial dysfunction, ERK1\\/2 inactivation, COX-2 down-regulation, and apoptosis. In conclusion, our results suggest that LY83583 may represent a novel therapeutic agent for the treatment of RCC, which remains highly refractory to antineoplastic agents. Our data provide a molecular basis for the anticancer activity of LY83583.

  6. Stable low-level expression of p21WAF1/CIP1 in A549 human bronchogenic carcinoma cell line-derived clones down-regulates E2F1 mRNA and restores cell proliferation control

    Directory of Open Access Journals (Sweden)

    Crawford Erin L

    2006-01-01

    Full Text Available Abstract Background Deregulated cell cycle progression and loss of proliferation control are key properties of malignant cells. In previous studies, an interactive transcript abundance index (ITAI comprising three cell cycle control genes, [MYC × E2F1]/p21 accurately distinguished normal from malignant bronchial epithelial cells (BEC, using a cut-off threshold of 7,000. This cut-off is represented by a line with a slope of 7,000 on a bivariate plot of p21 versus [MYC × E2F1], with malignant BEC above the line and normal BEC below the line. This study was an effort to better quantify, at the transcript abundance level, the difference between normal and malignant BEC. The hypothesis was tested that experimental elevation of p21 in a malignant BEC line would decrease the value of the [MYC × E2F1]/p21 ITAI to a level below this line, resulting in loss of immortality and limited cell population doubling capacity. In order to test the hypothesis, a p21 expression vector was transfected into the A549 human bronchogenic carcinoma cell line, which has low constitutive p21 TA expression relative to normal BEC. Results Following transfection of p21, four A549/p21 clones with stable two-fold up-regulated p21 expression were isolated and expanded. For each clone, the increase in p21 transcript abundance (TA was associated with increased total p21 protein level, more than 5-fold reduction in E2F1 TA, and 10-fold reduction in the [MYC × E2F1]/p21 ITAI to a value below the cut-off threshold. These changes in regulation of cell cycle control genes were associated with restoration of cell proliferation control. Specifically, each transfectant was capable of only 15 population doublings compared with unlimited population doublings for parental A549. This change was associated with an approximate 2-fold increase in population doubling time to 38.4 hours (from 22.3 hrs, resumption of contact-inhibition, and reduced dividing cell fraction as measured by flow

  7. TCR down-regulation controls T cell homeostasis

    DEFF Research Database (Denmark)

    Boding, Lasse; Bonefeld, Charlotte Menné; Nielsen, Bodil L

    2009-01-01

    TCR and cytokine receptor signaling play key roles in the complex homeostatic mechanisms that maintain a relative stable number of T cells throughout life. Despite the homeostatic mechanisms, a slow decline in naive T cells is typically observed with age. The CD3gamma di-leucine-based motif...... controls TCR down-regulation and plays a central role in fine-tuning TCR expression and signaling in T cells. In this study, we show that the age-associated decline of naive T cells is strongly accelerated in CD3gammaLLAA knock-in mice homozygous for a double leucine to alanine mutation in the CD3gamma di......-leucine-based motif, whereas the number of memory T cells is unaffected by the mutation. This results in premature T cell population senescence with a severe dominance of memory T cells and very few naive T cells in middle-aged to old CD3gamma mutant mice. The reduced number of naive T cells in CD3gamma mutant mice...

  8. miR-297 modulates multidrug resistance in human colorectal carcinoma by down-regulating MRP-2.

    Science.gov (United States)

    Xu, Ke; Liang, Xin; Shen, Ke; Cui, Daling; Zheng, Yuanhong; Xu, Jianhua; Fan, Zhongze; Qiu, Yanyan; Li, Qi; Ni, Lei; Liu, Jianwen

    2012-09-01

    Colorectal carcinoma is a frequent cause of cancer-related death in men and women. miRNAs (microRNAs) are endogenous small non-coding RNAs that regulate gene expression negatively at the post-transcriptional level. In the present study we investigated the possible role of microRNAs in the development of MDR (multidrug resistance) in colorectal carcinoma cells. We analysed miRNA expression levels between MDR colorectal carcinoma cell line HCT116/L-OHP cells and their parent cell line HCT116 using a miRNA microarray. miR-297 showed lower expression in HCT116/L-OHP cells compared with its parental cells. MRP-2 (MDR-associated protein 2) is an important MDR protein in platinum-drug-resistance cells and is a predicted target of miR-297. Additionally miR-297 was down-regulated in a panel of human colorectal carcinoma tissues and negatively correlated with expression levels of MRP-2. Furthermore, we found that ectopic expression of miR-297 in MDR colorectal carcinoma cells reduced MRP-2 protein level and sensitized these cells to anti-cancer drugs in vitro and in vivo. Taken together, our findings suggest that miR-297 could play a role in the development of MDR in colorectal carcinoma cells, at least in part by modulation of MRP-2.

  9. Expression of NDRG2 is down-regulated in high-risk adenomas and colorectal carcinoma

    DEFF Research Database (Denmark)

    Lorentzen, Anders; Vogel, Lotte K.; Lewinsky, Rikke H;

    2007-01-01

    BACKGROUND: It has recently been shown that NDRG2 mRNA is down-regulated or undetectable in several human cancers and cancer cell-lines. Although the function of NDRG2 is unknown, high NDRG2 expression correlates with improved prognosis in high-grade gliomas. The aim of this study has been to exa...

  10. Down-regulation of p73 correlates with high histological grade in Japanese with breast carcinomas

    Institute of Scientific and Technical Information of China (English)

    DU Cai-wen; Izo Kimijima; Toru Otake; Rikiya Abe; Seiichi Takenoshita; ZHANG Guo-jun

    2011-01-01

    Background p73, a homologue of p53, has been located at chromosome 1 p36-33, a region of frequently observed loss of heterozygosity in breast cancers. The objective of the present study was to investigate the function of p73 in Japanese with breast cancers. Methods Sixty Japanese patients with breast cancer were assessed by polymerase chain reaction single strand confirmation polymorphism analysis and direct sequencing to detect the p73 allele. p73 mRNA levels were also determined in 40 out of 60 patients by reverse-transcriptional polymerase chain reaction. Results We analyzed the entire open reading frame of the p73 gene by polymerase chain reaction single strand confirmation polymorphism and sequencing, and failed to identify any mutations of p73 in the encoding regions detected.Loss of heterozygosity of p73 was infrequent and only found in 9% of breast carcinomas. We revealed a few polymorphisms with a frequency of 13%-29%, which had been reported previously. Down-regulation of p73 mRNA expression was observed in tumor tissues in comparison to the normal breast tissues. A significant inverse correlation was found between p73 transcripts and high histological grade, suggesting that down-regulated p73 expression could be related to poor prognosis in those patients. Conclusion Our results suggest that p73 may serve as a tumor suppressor gene and its expression plays a role in tumorigenesis in Japanese patients with breast cancer.

  11. Hepatitis B Virus Down-Regulates Expressions of MHC Class ⅠMolecules on Hepatoplastoma Cell Line

    Institute of Scientific and Technical Information of China (English)

    Yongyan Chen; Min Cheng; ,Zhigang Tian

    2006-01-01

    Chronic HBV infection is associated with a 100-fold high risk of developing hepatocellular carcinoma. Tumor recognition is of the most importance during the immune surveillance process that prevents cancer development in humans. In the present study, the expressions of MHC class Ⅰ molecules on hepatoplastoma cell line HepG2.2.15 were investigated to indicate the possible effects of HBV on the immune recognition during HBV-associated hepatocellular carcinoma. It was found that the expressions of MHC class Ⅰ molecules HLA-ABC, HLA-E and MICA were much lower in HepG2.2.15 cells compared with HepG2 cells. The expressing HBV in human hepatoplastoma cell line significantly down-regulated the expressions of MHC class Ⅰ molecules. Additionally, it was observed that in murine chronic HBsAg carriers the expression of classical MHC-Ⅰ molecule on hepatocytes was down-regulated. These results demonstrated that HBV might affect the immune recognition during HBV-associated hepatoceilular carcinoma such as the recognition of CD8+ T, NK-CTL and NK cells and prevent the immune surveillance against tumors. However, the effects of HBV down-regulation of MHC class Ⅰ molecules on the target cells in vivo should be further studied.

  12. Copy number changes of target genes in chromosome 3q25.3-qter of esophageal squamous cell carcinoma: TP63 is amplified in early carcinogenesis but down-regulated as disease progressed

    Institute of Scientific and Technical Information of China (English)

    Chueh-Chuan Yen; Liang-Shun Wang; Min-Hsiung Huang; Biing-Shiung Huang; Cheng-Po Hu; Po-Min Chen; Chi-Hung Lin; Yann-Jang Chen; Chin-Chen Pan; Kai-Hsi Lu; Paul Chih-Hsueh Chen; Jiun-Yi Hsia; Jung-Ta Chen; Yu-Chung Wu; Wen-Hu Hsu

    2005-01-01

    AIM: By using comparative genomic hybridization, gain of 3q was found in 45-86% cases of esophageal squamous cell carcinoma (EC-SCC). Chromosome 3q25.3-qter is the minimal common region with several oncogenes found within this region. However, amplification patterns of these genes in EC-SCC have never been reported. The possible association of copy number changes of these genes with pathologic characteristics is still not clear.METHODS: Real-time quantitative PCR (Q-PCR) was performed to analyze the copy number changes of 13candidate genes within this region in 60 primary tumors of EC-SCC, and possible association of copy number changes with pathologic characteristics was analyzed by statistics. Immunohistochemistry (IHC) study was also performed on another set of 111 primary tumors of EC-SCC to verify the association between TP63 expression change and lymph node metastasis status.RESULTS: The average copy numbers (±SE) per haploid genome of individual genes in 60 samples were (from centromere to telomere): SSR3:4.19 (±0.69); CCNL1:5.24 (±0.67); SvC4L1: 2.01 (±0.16); EVI1: 2.02 (±0.12);hTERC: 5.28 (±0.54); SKIL: 2.71 (±0.14); EIF5A2: 1.95(±0.12); ECT2:9.18 (±1.68); PIK3CA: 8.13 (±1.17);EIF4G1:1.07 (±0.05); SST: 3.07 (±0.25); TP63: 2.51(±0.22); TFRC: 2.42 (±0.19). Four clusters of amplification were found: SSR3 and CCLN1 at 3q25.31; hTERC andSKIL at 3q26.2; ECT2 and PIK3CA at 3q26.31-q26.32; and SST, TP63 and TFRC at 3q27.3-q29. Patients with lymph node metastasis had significantly lower copy number of TP63 in the primary tumor than those without lymph node metastasis. IHC study on tissue arrays also showed that patients with lymph node metastasis have significantly lower TP63 staining score in the primary tumor than those without lymph node metastasis.CONCLUSION: This study showed that different amplification patterns were seen among different genes within 3q25.3-qter in EC-SCC, and several novel candidate oncogenes(SSR3, SMC4L1, ECT2, and SST) were

  13. Curcumin significantly enhances dual PI3K/Akt and mTOR inhibitor NVP-BEZ235-induced apoptosis in human renal carcinoma Caki cells through down-regulation of p53-dependent Bcl-2 expression and inhibition of Mcl-1 protein stability.

    Directory of Open Access Journals (Sweden)

    Bo Ram Seo

    Full Text Available The PI3K/Akt and mTOR signaling pathways are important for cell survival and growth, and they are highly activated in cancer cells compared with normal cells. Therefore, these signaling pathways are targets for inducing cancer cell death. The dual PI3K/Akt and mTOR inhibitor NVP-BEZ235 completely inhibited both signaling pathways. However, NVP-BEZ235 had no effect on cell death in human renal carcinoma Caki cells. We tested whether combined treatment with natural compounds and NVP-BEZ235 could induce cell death. Among several chemopreventive agents, curcumin, a natural biologically active compound that is extracted from the rhizomes of Curcuma species, markedly induced apoptosis in NVP-BEZ235-treated cells. Co-treatment with curcumin and NVP-BEZ235 led to the down-regulation of Mcl-1 protein expression but not mRNA expression. Ectopic expression of Mcl-1 completely inhibited curcumin plus NVP-NEZ235-induced apoptosis. Furthermore, the down-regulation of Bcl-2 was involved in curcumin plus NVP-BEZ235-induced apoptosis. Curcumin or NVP-BEZ235 alone did not change Bcl-2 mRNA or protein expression, but co-treatment reduced Bcl-2 mRNA and protein expression. Combined treatment with NVP-BEZ235 and curcumin reduced Bcl-2 expression in wild-type p53 HCT116 human colon carcinoma cells but not p53-null HCT116 cells. Moreover, Bcl-2 expression was completely reversed by treatment with pifithrin-α, a p53-specific inhibitor. Ectopic expression of Bcl-2 also inhibited apoptosis in NVP-BE235 plus curcumin-treated cells. In contrast, NVP-BEZ235 combined with curcumin did not have a synergistic effect on normal human skin fibroblasts and normal human mesangial cells. Taken together, combined treatment with NVP-BEZ235 and curcumin induces apoptosis through p53-dependent Bcl-2 mRNA down-regulation at the transcriptional level and Mcl-1 protein down-regulation at the post-transcriptional level.

  14. Neocytolysis: physiological down-regulator of red-cell mass

    Science.gov (United States)

    Alfrey, C. P.; Rice, L.; Udden, M. M.; Driscoll, T. B.

    1997-01-01

    It is usually considered that red-cell mass is controlled by erythropoietin-driven bone marrow red-cell production, and no physiological mechanisms can shorten survival of circulating red cells. In adapting to acute plethora in microgravity, astronauts' red-cell mass falls too rapidly to be explained by diminished red-cell production. Ferrokinetics show no early decline in erythropolesis, but red cells radiolabelled 12 days before launch survive normally. Selective destruction of the youngest circulating red cells-a process we call neocytolysis-is the only plausible explanation. A fall in erythropoietin below a threshold is likely to initiate neocytolysis, probably by influencing surface-adhesion molecules. Recognition of neocytolysis will require re-examination of the pathophysiology and treatment of several blood disorders, including the anaemia of renal disease.

  15. TCR down-regulation boosts T-cell-mediated cytotoxicity and protection against poxvirus infections

    DEFF Research Database (Denmark)

    Hansen, Ann Kathrine; Regner, Matthias; Bonefeld, Charlotte Menne

    2011-01-01

    Cytotoxic T (Tc) cells play a key role in the defense against virus infections. Tc cells recognize infected cells via the T-cell receptor (TCR) and subsequently kill the target cells by one or more cytotoxic mechanisms. Induction of the cytotoxic mechanisms is finely tuned by the activation signals...... from the TCR. To determine whether TCR down-regulation affects the cytotoxicity of Tc cells, we studied TCR down-regulation-deficient CD3¿LLAA mice. We found that Tc cells from CD3¿LLAA mice have reduced cytotoxicity due to a specific deficiency in exocytosis of lytic granules. To determine whether......-regulation critically increases Tc cell cytotoxicity and protection against poxvirus infection....

  16. Down-regulated expression of atypical PKC-binding domain deleted asip isoforms in human hepatocellular carcinomas

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Asip is a mammalian homologue of polarity protein Par-3 of Caenorhabditis elegans and Bazooka of Drosophila melanogaster. Asip/Par-3/Bazooka are PDZ-motif containing proteins that localize asymmetrically to the cell periphery and play a pivotal role in cell polarity and asymmetric cell division. In the present study, we have cloned human asip cDNA and its splicing variants by 5'-RACE and RT-PCR using candidate human EST clones which have a high homology to rat asip cDNA. The full-length cDNA of human asip encodes a 1,353 aa protein exhibiting 88% similarity to the rat one. Human asip is a single copy gene consisting of at least 26 exons and localizing in human chromosome 10, band p11.2, with some extraordinarily long introns. All exon/intron boundary nucleotides conform to the “gt-ag” rule. Three main transcripts were detected by Northern blot analysis, and at least five variants, from alternative splicing and polyadenylation, have been identified by RT-PCR and liver cDNA library screening. Exon 17b deleted asip mRNAs expressed ubiquitously in normal human tissues, including liver, on RT-PCR analysis. However, they were absent from most human liver cancer cell lines examined. More interestingly, the expression of exon 17b deleted variants was down regulated in 52.6% (10/19) clinic specimens of human hepatocellular carcinomas (HCCs), compared with the surrounding nontumorous liver tissues from the same patients. The presence of various splicing transcripts, the variation of their distribution among different tissues and cells, and their differential expressions in human HCCs suggest that human Asip isoforms may function in different context.

  17. LAPTM4B Down Regulation Inhibits the Proliferation, Invasion and Angiogenesis of HeLa Cells In Vitro

    Directory of Open Access Journals (Sweden)

    Fanling Meng

    2015-09-01

    Full Text Available Background/Aims: LAPTM4B (lysosome-associated protein transmembrane 4 beta is a novel oncogene with important functions in aggressive human carcinomas, including cervical cancer. However, the specific functions and internal molecular mechanisms associated with this gene in the context of cervical cancer remain unclear. Methods: In this study, we explored the effects and mechanisms of LAPTM4B on tumor growth, metastasis and angiogenesis in vitro by depletion of LAPTM4B in Hela cell. RNA interference was used to induce down regulation of LAPTM4B gene expression in Hela cells. The motility, migration potential, and proliferation of the Hela cells were measured by flow cytometry, Transwell migration assays, wound healing assays, and Cell Counting Kit-8 assays. In addition, the cell cycle analysis utilized fluorescence-activated cell sorting. Results: In this study, RNAi-mediated LAPTM4B knockdown inhibited cell growth and angiogenesis. In vitro, HeLa cells with down regulated LAPTM4B also exhibited decreased migration and invasion activity as well as significantly reduced CDK12, HIF-1α, MMP-2, MMP-9 and VEGF expression. LAPTM4B blockade significantly decreased cord lengths and branch points in a tube formation assay. Conclusions: These results suggested that LAPTM4B inactivation could be a novel therapeutic target for cervical cancer.

  18. SAMHD1 is down regulated in lung cancer by methylation and inhibits tumor cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Jia-lei [Department of Medical Oncology, Fudan University Shanghai Cancer Center, Shanghai 200032 (China); Lu, Fan-zhen [Department of Thoracic Surgery, The Huadong Hospital, Fudan University, Shanghai 200040 (China); Shen, Xiao-Yong, E-mail: shengxiaoyong_sh@163.com [Department of Thoracic Surgery, The Huadong Hospital, Fudan University, Shanghai 200040 (China); Wu, Yun, E-mail: WuYun_hd@163.com [Department of Thoracic Surgery, The Huadong Hospital, Fudan University, Shanghai 200040 (China); Zhao, Li-ting [Department of Thoracic Surgery, The Huadong Hospital, Fudan University, Shanghai 200040 (China)

    2014-12-12

    Highlights: • SAMHD1 expression level is down regulated in lung adenocarcinoma. • The promoter of SAMHD1 is methylated in lung adenocarcinoma. • Over expression of SAMHD1 inhibits the proliferation of lung cancer cells. - Abstract: The function of dNTP hydrolase SAMHD1 as a viral restriction factor to inhibit the replication of several viruses in human immune cells was well established. However, its regulation and function in lung cancer have been elusive. Here, we report that SAMHD1 is down regulated both on protein and mRNA levels in lung adenocarcinoma compared to adjacent normal tissue. We also found that SAMHD1 promoter is highly methylated in lung adenocarcinoma, which may inhibit its gene expression. Furthermore, over expression of the SAMHD1 reduces dNTP level and inhibits the proliferation of lung tumor cells. These results reveal the regulation and function of SAMHD1 in lung cancer, which is important for the proliferation of lung tumor cells.

  19. Down regulation of ribosomal protein mRNAs during neuronal differentiation of human NTERA2 cells.

    Science.gov (United States)

    Bévort, M; Leffers, H

    2000-10-01

    We have analysed the expression of 32 ribosomal protein (RP) mRNAs during retinoic acid induced neuronal differentiation of human NTERA2 cells. Except for a new S27 variant (S27v), all were down regulated both in selectively replated differentiated neurons and the most differentiated continuous cultures, i.e., non-replated cultures. However, the expression profiles of the individual RP mRNAs were different, most (L3, L7, L8, L10, L13, L23a, L27a, L36a, L39, P0, S2, S3, S3a, S4X, S6, S9, S12, S13, S16, S19, S20, S23, and S27a) exhibited a constant down regulation, whereas a few were either initially constant (L11, L32, S8, and S11) or up regulated (L6, L15, L17, L31, and S27y) and then down regulated. The expression of S27v remained elevated in the most differentiated continuous cultures but was down regulated in replated differentiated neurons. The down regulation of RP mRNAs was variable: the expression levels in differentiated replated neurons were between 10% (S3) and 90% (S11) of the levels in undifferentiated cells. The ratio between rRNA and RP mRNA changed during the differentiation; in differentiated neurons there were, on average, about half the number of RP mRNAs per rRNA as compared to undifferentiated cells. The expression profiles of a few translation-related proteins were also determined. EF1alpha1, EF1beta1, and EF1delta were down regulated, whereas the expression of the neuron and muscle specific EF1alpha2 increased. The reduction in the expression of RP mRNAs was coordinated with a reduction in the expression level of the proliferation marker PCNA. The expression levels of most RP mRNAs were lower in purified differentiated post-mitotic neurons than in the most differentiated continuous cultures, despite similar levels of PCNA, suggesting that both the differentiation state and the proliferative status of the cells affect the expression of RP mRNAs.

  20. TCR Down-Regulation Controls Virus-Specific CD8+ T Cell Responses

    DEFF Research Database (Denmark)

    Bonefeld, Charlotte Menné; Haks, Mariëlle; Nielsen, Bodil

    2008-01-01

    The CD3gamma di-leucine-based motif plays a central role in TCR down-regulation. However, little is understood about the role of the CD3gamma di-leucine-based motif in physiological T cell responses. In this study, we show that the expansion in numbers of virus-specific CD8(+) T cells is impaired...... in mice with a mutated CD3gamma di-leucine-based motif. The CD3gamma mutation did not impair early TCR signaling, nor did it compromise recruitment or proliferation of virus-specific T cells, but it increased the apoptosis rate of the activated T cells by increasing down-regulation of the antiapoptotic...... molecule Bcl-2. This resulted in a 2-fold reduction in the clonal expansion of virus-specific CD8(+) T cells during the acute phase of vesicular stomatitis virus and lymphocytic choriomeningitis virus infections. These results identify an important role of CD3gamma-mediated TCR down-regulation in virus...

  1. Trop-2表达下调对食管鳞癌EC9706细胞增殖和细胞迁移的影响%Down-regulation of Trop-2 expression inhibits cell proliferation and migration in human esophageal squamous cell carcinoma cell line EC9706

    Institute of Scientific and Technical Information of China (English)

    李慎柯; 范天黎; 寇洁; 崔瑶

    2011-01-01

    目的:研究肿瘤相关钙信号传导蛋白2(tumorassociated calcium signal transducer-2,Trop-2)表达下调对食管鳞癌EC9706细胞增殖和细胞迁移的影响,并探讨其可能的分子机制.方法:将Trop-2 siRNA和对照siRNA转染食管鳞癌EC9706细胞,利用实时荧光定量PCR和Western blotting检测转染Trop-2 siRNA后食管鳞癌EC9706细胞中Trop-2 mRNA和蛋白的表达.利用CCK-8试剂分析转染Trop-2 siRNA后EC9706细胞的增殖变化,接着采用Boyden小室研究下调Trop-2对EC9706细胞迁移的影响,最后利用Western blotting技术分析与细胞迁移相关基因MMP-7表达的变化.结果:转染Trop-2 48 h,Trop-2 siRNA组中Trop-2 mRNA和蛋白的表达显著低于未处理组和对照siRNA组(P<0.05).Trop-2表达下调能明显抑制食管鳞癌EC9706细胞的增殖(P<0.05).此外,Boyden小室结果表明,与未处理组(119.27±4.63)及对照siRNA组(112.81±5.01)相比,Trop-2 siRNA组中EC9706细胞的穿膜数(65.29±4.33)明显降低.Westernblotting结果表明,Trop-2 siRNA能明显降低食管鳞癌EC9706细胞中MMP-7的表达.结论:Trop-2表达下调能抑制食管鳞癌细胞的增殖和迁移,其介导的迁移能力的降低可能与MMP-7表达的下调密切相关,操纵Trop-2有望成为食管鳞癌的分子治疗靶点.%AIM: To investigate the effect of down-regulation of tumor-associated calcium signal trans-ducer-2 (Trop-2) expression on cell proliferation and migration in human esophageal squamous cell carcinoma (ESCC cell line EC9706 and to explore possible molecular mechanisms involved.METHODS: Trop-2-specific small interferingRNA (siRNA) and control siRNA were transfect-ed into EC9706 cells, and expression of Trop-2 mRNA and protein was detected by real-time PCR and Western blotting. CCK-8 assay was used to measure cell proliferation, and Boyden chamber assay was utilized to determine cell migration. The expression of cell migration-related MMP-7 protein was detected by Western blotting

  2. [BMMSC from blastic phase CML down-regulate leukemia cell apoptosis].

    Science.gov (United States)

    Wang, Ying; Han, Yu-Xiang; Niu, Zhi-Yun; Wang, Xing-Zhe; Hua, Huan; Shang, Yin-Tao; Wang, Fu-Xu; Zhang, Xue-Jun; Luo, Jian-Min

    2014-10-01

    The purpose of this study was to investigate the effect of bone marrow mesenchymal stem cells (BMMSC) from patients with chronic myeloid leukemia (CML) in blastic phase (Bp) on K562 cells and the primary CML-Bp cells, and to explore its potential mechanisms. K562 cells and primary CML-Bp cells were co-cultured with BMMSC of different groups; the cell proliferation was detected by MTT method, the cell apoptosis rate and mitochondrial membrane potential were measured by flow cytometry, the expression levels of Caspase-8, Caspase-9, and activated Caspase-3 in cells were measured by Western blot. The results showed that the CML-Bp BMMSC could enhance the survival rate of K562 cells treated with adviamycin (ADM) and display protective effect on K562 cells and primary CML-Bp mononuctear cells, inhibited ADM-induced leukimia cell apoptosis (P < 0.05); as compared with CML-chronic phase (CML-Cp) BMMSC and normal BMMSC, the CML-Bp BMMSC showed the highest protective effect on leukemic cells, the mitochondrial membrane potential of co-cultured cells slightly droped (P < 0.05). In the CML-Bp BMMSC cultured with K562 cells, the expression level of caspase-3 was more down-regulated than that in K562 alone plus ADM group, while the expression of caspase-9 significantly increased (P < 0.05). It is concluded that the CML-Bp BMMSC down-regulates ADM-induced leukemia cell appoptosis, its mechanism may relate with the inhibition of mitochondrial membrane potential drop, the stabilization of unactive expression of caspase-9 and down-regulation of caspase-3 expression.

  3. Lupeol, a dietary triterpene, inhibited growth, and induced apoptosis through down-regulation of DR3 in SMMC7721 cells.

    Science.gov (United States)

    Zhang, Lin; Zhang, Youcheng; Zhang, Lingyi; Yang, Xiaojun; Lv, Zhicheng

    2009-02-01

    Lupeol (Lup-20(29)-en-3H-ol), a novel dietary triterpene, was found in fruits, vegetables, and several medicinal plants. Here, we investigated its growth-inhibitory effect and associated mechanisms in hepatocellular carcinoma SMMC7721 cells. Lupeol treatment resulted in significant inhibition of cell viability in a dose-dependent manner and caused apoptotic death of this cell line with activation of caspase3 expression. Caspase8 inhibitor pretreatment was found to partially block the apoptosis induced by Lupeol. Moreover, Lupeol specifically caused a significant decrease in the expression of Death receptor 3 (DR3) mRNA and protein and a significant elevated expression of FADD mRNA whereas Fas mRNA and protein expression was not detectable. Further more, knockdown of DR3 by small interfering RNA inhibited the growth and induced apoptosis of hepatocellular carcinoma cell. These results suggested that Lupeol treatment induced growth inhibition and apoptosis in SMMC7721 cells, the mechanism is due to down-regulation of DR3 expression. We demonstrated that Lupeol appears to be a promising chemopreventive agent for treating hepatocellular carcinoma, and DR3 may be an important target for liver cancer therapy.

  4. CHIP mediates down-regulation of nucleobindin-1 in preosteoblast cell line models.

    Science.gov (United States)

    Xue, Fuying; Wu, Yanping; Zhao, Xinghui; Zhao, Taoran; Meng, Ying; Zhao, Zhanzhong; Guo, Junwei; Chen, Wei

    2016-08-01

    Nucleobindin-1 (NUCB1), also known as Calnuc, is a highly conserved, multifunctional protein widely expressed in tissues and cells. It contains two EF-hand motifs which have been shown to play a crucial role in binding Ca(2+) ions. In this study, we applied comparative two-dimensional gel electrophoresis to characterize differentially expressed proteins in HA-CHIP over-expressed and endogenous CHIP depleted MC3T3-E1 stable cell lines, identifying NUCB1 as a novel CHIP/Stub1 targeted protein. NUCB1 interacts with and is down-regulated by CHIP by both proteasomal dependent and independent pathways, suggesting that CHIP-mediated down-regulation of nucleobindin-1 might play a role in osteoblast differentiation. The chaperone protein Hsp70 was found to be important for CHIP and NUCB1 interaction as well as CHIP-mediated NUCB1 down-regulation. Our findings provide new insights into understanding the stability regulation of NUCB1.

  5. Effects of p21 Gene Down-Regulation through RNAi on Antler Stem Cells In Vitro.

    Directory of Open Access Journals (Sweden)

    Qianqian Guo

    Full Text Available Cell cycle is an integral part of cell proliferation, and consists mainly of four phases, G1, S, G2 and M. The p21 protein, a cyclin dependent kinase inhibitor, plays a key role in regulating cell cyclevia G1 phase control. Cells capable of epimorphic regeneration have G2/M accumulation as their distinctive feature, whilst the majority of somatic cells rest at G1 phase. To investigate the role played byp21 in antler regeneration, we studied the cell cycle distribution of antler stem cells (ASCs, via down-regulation of p21 in vitro using RNAi. The results showed that ASCs had high levels of p21 mRNA expression and rested at G1 phase, which was comparable to the control somatic cells. Down-regulation of p21 did not result in ASC cell cycle re-distribution toward G2/M accumulation, but DNA damage and apoptosis of the ASCs significantly increased and the process of cell aging was slowed. These findings suggest that the ASCs may have evolved to use an alternative, p21-independent cell cycle regulation mechanism. Also a unique p21-dependent inhibitory effect may control DNA damage as a protective mechanism to ensure the fast proliferating ASCs do not become dysplastic/cancerous. Understanding of the mechanism underlying the role played by p21 in the ASCs could give insight into a mammalian system where epimorphic regeneration is initiated whilst the genome stability is effectively maintained.

  6. Effects of p21 Gene Down-Regulation through RNAi on Antler Stem Cells In Vitro

    Science.gov (United States)

    Guo, Qianqian; Wang, Datao; Liu, Zhen; Li, Chunyi

    2015-01-01

    Cell cycle is an integral part of cell proliferation, and consists mainly of four phases, G1, S, G2 and M. The p21 protein, a cyclin dependent kinase inhibitor, plays a key role in regulating cell cyclevia G1 phase control. Cells capable of epimorphic regeneration have G2/M accumulation as their distinctive feature, whilst the majority of somatic cells rest at G1 phase. To investigate the role played byp21 in antler regeneration, we studied the cell cycle distribution of antler stem cells (ASCs), via down-regulation of p21 in vitro using RNAi. The results showed that ASCs had high levels of p21 mRNA expression and rested at G1 phase, which was comparable to the control somatic cells. Down-regulation of p21 did not result in ASC cell cycle re-distribution toward G2/M accumulation, but DNA damage and apoptosis of the ASCs significantly increased and the process of cell aging was slowed. These findings suggest that the ASCs may have evolved to use an alternative, p21-independent cell cycle regulation mechanism. Also a unique p21-dependent inhibitory effect may control DNA damage as a protective mechanism to ensure the fast proliferating ASCs do not become dysplastic/cancerous. Understanding of the mechanism underlying the role played by p21 in the ASCs could give insight into a mammalian system where epimorphic regeneration is initiated whilst the genome stability is effectively maintained. PMID:26308075

  7. Effects of p21 Gene Down-Regulation through RNAi on Antler Stem Cells In Vitro.

    Science.gov (United States)

    Guo, Qianqian; Wang, Datao; Liu, Zhen; Li, Chunyi

    2015-01-01

    Cell cycle is an integral part of cell proliferation, and consists mainly of four phases, G1, S, G2 and M. The p21 protein, a cyclin dependent kinase inhibitor, plays a key role in regulating cell cyclevia G1 phase control. Cells capable of epimorphic regeneration have G2/M accumulation as their distinctive feature, whilst the majority of somatic cells rest at G1 phase. To investigate the role played byp21 in antler regeneration, we studied the cell cycle distribution of antler stem cells (ASCs), via down-regulation of p21 in vitro using RNAi. The results showed that ASCs had high levels of p21 mRNA expression and rested at G1 phase, which was comparable to the control somatic cells. Down-regulation of p21 did not result in ASC cell cycle re-distribution toward G2/M accumulation, but DNA damage and apoptosis of the ASCs significantly increased and the process of cell aging was slowed. These findings suggest that the ASCs may have evolved to use an alternative, p21-independent cell cycle regulation mechanism. Also a unique p21-dependent inhibitory effect may control DNA damage as a protective mechanism to ensure the fast proliferating ASCs do not become dysplastic/cancerous. Understanding of the mechanism underlying the role played by p21 in the ASCs could give insight into a mammalian system where epimorphic regeneration is initiated whilst the genome stability is effectively maintained.

  8. Down-regulation of Rab5 decreases characteristics associated with maintenance of cell transformation

    Energy Technology Data Exchange (ETDEWEB)

    Silva, Patricio; Soto, Nicolás [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Díaz, Jorge [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Center for Molecular Studies of the Cell, Institute of Biomedical Sciences (ICBM), Faculty of Medicine, Universidad de Chile, Santiago (Chile); Mendoza, Pablo [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Díaz, Natalia [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Center for Molecular Studies of the Cell, Institute of Biomedical Sciences (ICBM), Faculty of Medicine, Universidad de Chile, Santiago (Chile); Quest, Andrew F.G. [Center for Molecular Studies of the Cell, Institute of Biomedical Sciences (ICBM), Faculty of Medicine, Universidad de Chile, Santiago (Chile); Advanced Center for Chronic Diseases (ACCDiS), Faculty of Medicine, Universidad de Chile, Santiago (Chile); Torres, Vicente A., E-mail: vatorres@med.uchile.cl [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Advanced Center for Chronic Diseases (ACCDiS), Faculty of Medicine, Universidad de Chile, Santiago (Chile)

    2015-08-21

    The early endosomal protein Rab5 is highly expressed in tumor samples, although a causal relationship between Rab5 expression and cell transformation has not been established. Here, we report the functional effects of targeting endogenous Rab5 with specific shRNA sequences in different tumor cell lines. Rab5 down-regulation in B16-F10 cells decreased tumor formation by subcutaneous injection into C57/BL6 mice. Accordingly, Rab5 targeting in B16-F10 and A549, but not MDA-MB-231 cells was followed by decreased cell proliferation, increased apoptosis and decreased anchorage-independent growth. These findings suggest that Rab5 expression is required to maintain characteristics associated with cell transformation. - Highlights: • Rab5 is important to the maintenance of cell transformation characteristics. • Down-regulation of Rab5 decreases cell proliferation and increases apoptosis in different cancer cells. • Rab5 is required for anchorage-independent growth and tumorigenicity in-vivo.

  9. Androgen Depletion Induces Senescence in Prostate Cancer Cells through Down-regulation of Skp2

    Directory of Open Access Journals (Sweden)

    Zuzana Pernicová

    2011-06-01

    Full Text Available Although the induction of senescence in cancer cells is a potent mechanism of tumor suppression, senescent cells remain metabolically active and may secrete a broad spectrum of factors that promote tumorigenicity in neighboring malignant cells. Here we show that androgen deprivation therapy (ADT, a widely used treatment for advanced prostate cancer, induces a senescence-associated secretory phenotype in prostate cancer epithelial cells, indicated by increases in senescence-associated β-galactosidase activity, heterochromatin protein 1β foci, and expression of cathepsin B and insulin-like growth factor binding protein 3. Interestingly, ADT also induced high levels of vimentin expression in prostate cancer cell lines in vitro and in human prostate tumors in vivo. The induction of the senescence-associated secretory phenotype by androgen depletion was mediated, at least in part, by down-regulation of S-phase kinase-associated protein 2, whereas the neuroendocrine differentiation of prostate cancer cells was under separate control. These data demonstrate a previously unrecognized link between inhibition of androgen receptor signaling, down-regulation of S-phase kinase-associated protein 2, and the appearance of secretory, tumor-promoting senescent cells in prostate tumors. We propose that ADT may contribute to the development of androgen-independent prostate cancer through modulation of the tissue microenvironment by senescent cells.

  10. Down-regulation of ALKBH2 increases cisplatin sensitivity in H1299 lung cancer cells

    Institute of Scientific and Technical Information of China (English)

    Shuang-shuang WU; Wei XU; Shan LIU; Bo CHEN; Xue-li WANG; Yan WANG; Shi-feng LIU; Jian-qing WU

    2011-01-01

    Aim: To elucidate the combined effect of alkylated DNA repair protein alkB homolog 2 (ALKBH2)-targeting gene therapy and cisplatin (cDDP) chemotherapy on the non-small cell lung cancer (NSCLC)H1299 cell line.Methods: ALKBH2 was down-regulated in H1299 cells by lentivirus-mediated RNA interference (RNAi). Changes in ALKBH2 expression were determined using real-time RT-PCR and Western blotting. Cell viability was evaluated using MTT assay. DNA synthesis in proliferating cells was determined using BrdU incorporation assay. Cell apoptosis was determined using flow cytometry.Results: Lentivirus-mediated ALKBH2 silencing alone did not induce apoptosis or attenuate the growth potential of H1299 cells within five days post-infection. Combined treatment modalities with lentivirus-mediated ALKBH2 down-regulation and cDDP (333 μmol/L)were significantly more potent in inhibiting cell growth and inducing apoptosis than mono-chemotherapy.Conclusion: Combined treatment modalities of ALKBH2 knockdown and cDDP chemotherapy have the potential to improve the efficacy in the treatment of NSCLC.

  11. Glucosamine Modulates T Cell Differentiation through Down-regulating N-Linked Glycosylation of CD25.

    Science.gov (United States)

    Chien, Ming-Wei; Lin, Ming-Hong; Huang, Shing-Hwa; Fu, Shin-Huei; Hsu, Chao-Yuan; Yen, B Lin-Ju; Chen, Jiann-Torng; Chang, Deh-Ming; Sytwu, Huey-Kang

    2015-12-04

    Glucosamine has immunomodulatory effects on autoimmune diseases. However, the mechanism(s) through which glucosamine modulates different T cell subsets and diseases remain unclear. We demonstrate that glucosamine impedes Th1, Th2, and iTreg but promotes Th17 differentiation through down-regulating N-linked glycosylation of CD25 and subsequently inhibiting its downstream Stat5 signaling in a dose-dependent manner. The effect of glucosamine on T helper cell differentiation was similar to that induced by anti-IL-2 treatment, further supporting an IL-2 signaling-dependent modulation. Interestingly, excess glucose rescued this glucosamine-mediated regulation, suggesting a functional competition between glucose and glucosamine. High-dose glucosamine significantly decreased Glut1 N-glycosylation in Th1-polarized cells. This finding suggests that both down-regulated IL-2 signaling and Glut1-dependent glycolytic metabolism contribute to the inhibition of Th1 differentiation by glucosamine. Finally, glucosamine treatment inhibited Th1 cells in vivo, prolonged the survival of islet grafts in diabetic recipients, and exacerbated the severity of EAE. Taken together, our results indicate that glucosamine interferes with N-glycosylation of CD25, and thereby attenuates IL-2 downstream signaling. These effects suggest that glucosamine may be an important modulator of T cell differentiation and immune homeostasis.

  12. microRNA-143 down-regulates Hexokinase 2 in colon cancer cells

    DEFF Research Database (Denmark)

    Gregersen, Lea Haarup; Jacobsen, Anders; Frankel, Lisa;

    2012-01-01

    -regulated in several cancers including colon cancer. METHODS: To gain insight into the role of miR-143 in colon cancer, we used a microarray-based approach in combination with seed site enrichment analysis to identify miR-143 targets. RESULTS: As expected, transcripts down-regulated upon miR-143 overexpression had...... a significant enrichment of miR-143 seed sites in their 3' UTRs. Here we report the identification of Hexokinase 2 (HK2) as a direct target of miR-143. We show that re-introduction of miR-143 in the colon cancer cell line DLD-1 results in a decreased lactate secretion. CONCLUSION: We have identified...... and validated HK2 as a miR-143 target. Furthermore, our results indicate that miR-143 mediated down-regulation of HK2 affects glucose metabolism in colon cancer cells. We hypothesize that loss of miR-143-mediated repression of HK2 can promote glucose metabolism in cancer cells, contributing to the shift towards...

  13. Down-regulation of the DNA-repair endonuclease 8-oxo-guanine DNA glycosylase 1 (hOGG1) by sodium dichromate in cultured human A549 lung carcinoma cells.

    Science.gov (United States)

    Hodges, N J; Chipman, J K

    2002-01-01

    Hexavalent chromium is a genotoxic human pulmonary carcinogen that elevates DNA oxidation, apparently through the generation of reactive DNA-damaging intermediates including Cr(V), Cr(IV) and reactive oxygen species. We tested the hypothesis that elevation of DNA oxidation may also be through inhibition of the expression of the repair glycosylase for 8-oxo deoxyguanine (hOGG1) in cultured A549 human lung epithelial cells. Treatment with sodium dichromate (0-100 microM, 16 h) resulted in a concentration-dependent decrease in the levels of OGG1 mRNA as measured by both RT-PCR and RNase protection assay. Sodium dichromate at 25 microM and above gave a marked reduction of OGG1 mRNA expression which was not seen at 1 microM and below. No effect on the expression of the apurinic endonuclease hAPE or the house-keeping gene GAPDH was observed at any of the concentrations of sodium dichromate investigated. Treatment of cells with the pro-oxidant H(2)O(2) (0-200 microM) for 16 h had no detectable effect on the levels of OGG1 mRNA or protein expression suggesting that the effect of sodium dichromate is not mediated by H(2)O(2). Western blotting demonstrated that sodium dichromate (100 microM; 16 h and >25 microM; 28 h) markedly reduced levels of OGG1 protein in nuclear cell extracts. Additionally, treatment of cells with sodium dichromate (>25 microM, 28 h) resulted in a concentration-dependent decrease in the ability of nuclear extracts to nick a synthetic oligonucleotide containing 8-oxo deoxyguanine (8-oxo dG). We conclude that the elevation of 8-oxo dG levels observed in A549 cells treated with sodium dichromate may be, at least in part, due to a reduced capacity to repair endogenous and hexavalent chromium-induced 8-oxo dG.

  14. ATM down-regulation is associated with poor prognosis in sporadic breast carcinomas

    DEFF Research Database (Denmark)

    Bueno, R C; Canevari, R A; Villacis, R A R

    2014-01-01

    hybridization (aCGH) in 42 tumours. RESULTS: Low ATM levels were associated with tumour grade. Absence of ATM protein expression was associated with distant metastasis (P survival (DFS, P cancer-specific survival (CSS, P analysis indicated ATM......BACKGROUND: Ataxia telangiectasia-mutated (ATM) gene downexpression has been reported in sporadic breast carcinomas (BC); however, the prognostic value and mechanisms of ATM deregulation remain unclear. PATIENTS AND METHODS: ATM and miRNAs (miR-26a, miR-26b, miR-203, miR-421, miR-664, miR-576-5p...... and miR-18a) expression levels were evaluated by quantitative real-time PCR (RT-qPCR) in 52 BC and 3 normal breast samples. ATM protein expression was assessed by immunohistochemistry in 968 BC and 35 adjacent normal breast tissues. ATM copy number alteration was detected by array comparative genomic...

  15. Down-Regulation of Bcl-2 Protein Sensitizes NCI 460 Cells to Radiotherapy-Induced Apoptosis

    Institute of Scientific and Technical Information of China (English)

    Dongmei He; Yuan Zhang; Gexiu Liu

    2006-01-01

    OBJECTIVE To determine whether Bcl-2 protein down-regulation can render NCI-460 cells more susceptible to gamma radiation-induced apoptosis by treatment with antisense oligonucleotide (ASODN) against the coding region of Bcl-2 mRNA.METHODS Cell survival was determined using the trypan blue dye exclusion. Expression of the Bcl-2 protein was assayed using immunofluorescence labeling with fluoresce isothiocyanate. Apoptosis was determined by Giemsa staining and flow cytomertry.RESULTS It was found that Bcl-2 ASODN combined with radiation significantly reduced the number of viable cells (P<0.05). There was no difference in cell survival between a nonsense oligodeoxynucleotide/radiation combination and cells treated with radiation alone. Bcl-2 ASODN combined with radiation significantly inhibited expression of the Bcl-2protein in the NCI-H460 cells (P<0.05). Using Giemsa staining, cells treated with Bcl-2 ASODN combined with radiation at 72 h displayed classic apoptotic changes. Apoptotic rates of the NCI-H460 cells treated with Bcl-2 ASODN combined with radiation significantly increased (P<0.05), compared with either a nonsense oligodeoxynucleotide/radiation combination or radiation-treatment cells alone.CONCLUSION ASODN against the coding region of Bcl-2 mRNA increases radiation-induced apoptosis in NCI-H460 cells.

  16. MARCH1 down-regulation in IL-10-activated B cells increases MHC class II expression.

    Science.gov (United States)

    Galbas, Tristan; Steimle, Viktor; Lapointe, Réjean; Ishido, Satoshi; Thibodeau, Jacques

    2012-07-01

    IL-10 is vastly studied for its anti-inflammatory properties on most immune cells. However, it has been reported that IL-10 activates B cells, up-regulates their MHC class II molecules and prevents apoptosis. As MARCH1 was shown to be responsible for the intracellular sequestration of MHC class II molecules in dendritic cells and monocytes in response to IL-10, we set out to clarify the role of this ubiquitin ligase in B cells. Here, we demonstrate in mice that splenic follicular B cells represent the major cell population that up-regulate MHC II molecules in the presence of IL-10. Activation of these cells through TLR4, CD40 or the IL-10 receptor caused the down-regulation of MARCH1 mRNA. Accordingly, B cells from MARCH1-deficient mice do not up-regulate I-A(b) in response to IL-10. In all, our results demonstrate that IL-10 can have opposite effects on MARCH1 regulation in different cell types.

  17. Role of calcium signaling in down-regulation of aggrecan induced by cyclic tensile strain in annulus fibrosus cells

    Institute of Scientific and Technical Information of China (English)

    GUO Zhi-liang; ZHOU Yue; LI Hua-zhuang; CAO Guo-yong; TENG Hai-jun

    2006-01-01

    Objective:To study the role of intracellular calcium signal pathway in the down-regulation of aggrecan induced by cyclic tensile strain in the annulus fibrosus cells. Methods :The expression of aggrecan mRNA and core protein were respectively detected with RT-PCR and western blot after the channels transmitting calcium ions were blocked with EGTA, gadolinium and verapamil. Results:EGTA, gadolinium and verapamil partially prevented the effects of cyclic tensile strain on the expression of aggrecan in annulus fibrosus cells. Conclusion:The calcium signaling is involved in the down-regulation of proteoglycan resulting from cyclic tensile strain in the annulus fibrosus cells.

  18. Mechanism of Wnt signaling induced down regulation of mrhl long non-coding RNA in mouse spermatogonial cells

    Science.gov (United States)

    Akhade, Vijay Suresh; Dighe, Shrinivas Nivrutti; Kataruka, Shubhangini; Rao, Manchanahalli R. Satyanarayana

    2016-01-01

    Long non coding RNAs (lncRNAs) have emerged as important regulators of various biological processes. LncRNAs also behave as response elements or targets of signaling pathway(s) mediating cellular function. Wnt signaling is important in regulating mammalian spermatogenesis. Mrhl RNA negatively regulates canonical Wnt pathway and gets down regulated upon Wnt signaling activation in mouse spermatogonial cells. Also, mrhl RNA regulates expression of genes pertaining to Wnt pathway and spermatogenesis by binding to chromatin. In the present study, we delineate the detailed molecular mechanism of Wnt signaling induced mrhl RNA down regulation in mouse spermatogonial cells. Mrhl RNA has an independent transcription unit and our various experiments like Chromatin Immunoprecipitation (in cell line as well as mouse testis) and shRNA mediated down regulation convincingly show that β-catenin and TCF4, which are the key effector proteins of the Wnt signaling pathway are required for down regulation of mrhl RNA. We have identified Ctbp1 as the co-repressor and its occupancy on mrhl RNA promoter depends on both β-catenin and TCF4. Upon Wnt signaling activation, Ctbp1 mediated histone repression marks increase at the mrhl RNA promoter. We also demonstrate that Wnt signaling induced mrhl RNA down regulation results in an up regulation of various meiotic differentiation marker genes. PMID:26446991

  19. Down-Regulation of NDUFB9 Promotes Breast Cancer Cell Proliferation, Metastasis by Mediating Mitochondrial Metabolism.

    Directory of Open Access Journals (Sweden)

    Liang-Dong Li

    Full Text Available Despite advances in basic and clinical research, metastasis remains the leading cause of death in breast cancer patients. Genetic abnormalities in mitochondria, including mutations affecting complex I and oxidative phosphorylation, are found in breast cancers and might facilitate metastasis. Genes encoding complex I components have significant breast cancer prognostic value. In this study, we used quantitative proteomic analyses to compare a highly metastatic cancer cell line and a parental breast cancer cell line; and observed that NDUFB9, an accessory subunit of the mitochondrial membrane respiratory chain NADH dehydrogenase (complex I, was down-regulated in highly metastatic breast cancer cells. Furthermore, we demonstrated that loss of NDUFB9 promotes MDA-MB-231 cells proliferation, migration, and invasion because of elevated levels of mtROS, disturbance of the NAD+/NADH balance, and depletion of mtDNA. We also showed that, the Akt/mTOR/p70S6K signaling pathway and EMT might be involved in this mechanism. Thus, our findings contribute novel data to support the hypothesis that misregulation of mitochondrial complex I NADH dehydrogenase activity can profoundly enhance the aggressiveness of human breast cancer cells, suggesting that complex I deficiency is a potential and important biomarker for further basic research or clinical application.

  20. GPR30下调对子宫内膜癌细胞及裸鼠移植瘤组织 PI3K/Akt信号通路的影响%Influence of down-regulation of GPR30 on PI3 K/Akt signaling pathway in endometrial carcinoma cells and tumor tissue of nude mice

    Institute of Scientific and Technical Information of China (English)

    雷冬梅; 郭瑞霞; 刘泇希; 葛新; 乔玉环

    2015-01-01

    目的:探讨G蛋白偶联受体(GPR 30)下调对子宫内膜癌细胞及裸鼠移植瘤组织PI3K/Akt信号通路的影响。方法:采用免疫细胞化学SP法观察子宫内膜癌细胞系HEC-1A和Ishikawa细胞中GPR30、Akt和磷酸化Akt (p-Akt)蛋白的定位。以pGFP-V-RS(对照)和pGFP-V-RS-GPR30(干扰)分别转染两种细胞,用蛋白印迹法检测两种细胞中GPR30、Akt及p-Akt蛋白的表达水平。构建裸鼠移植瘤模型(分别接种上述4种转染细胞,共4组),用免疫组化SP法检测4组裸鼠移植瘤组织中p-Akt蛋白的表达。结果:①HEC-1A和Ishikawa细胞中,GPR30、Akt和p-Akt蛋白阳性表达均呈棕黄色,定位于细胞质。②稳定转染pGFP-VR-S -GPR30的两种细胞株中GPR30和p -Akt的表达均下调( P<0.05)。③与接种转染pGFP-V-RS细胞的2组裸鼠相比,接种转染pGFP-V-RS-GPR30细胞的2组裸鼠移植瘤组织中p-Akt表达降低。结论:在子宫内膜癌细胞及裸鼠移植瘤组织中, GPR30下调可能抑制了PI3K/Akt通路的活化。%Aim:To investigate the influence of down-regulation of G protein-coupled estrogen receptor 30 ( GPR30 ) on activation of PI3K/Akt signaling pathway in endometrial carcinoma cells and tumor tissue of nude mice .Methods:The location of GPR30, Akt and p-Akt protein in HEC-1A and Ishikawa cells was detected by immunocytochemical SP method . The expression of GPR30 in HEC-1A and Ishikawa cells was down-regulated by transfection with pGFP-V-RS-GPR30, a GPR30 antisense expression vector , and the cells transfected with pGFP-V-RS were the control; the levels of GPR30,Akt and p-Akt were detected by Western blot .The nude mice were allocated into 4 groups and inoculated the above transfected cells, and immunohistochemistry was performed to observe the changes of the expression level of p -Akt in the xenograft tis-sue.Results:Immunocytochemical SP method showed that GPR 30, Akt and p-Akt was stained as brown and

  1. microRNA-143 down-regulates Hexokinase 2 in colon cancer cells

    DEFF Research Database (Denmark)

    Gregersen, Lea Haarup; Jacobsen, Anders; Frankel, Lisa

    2012-01-01

    a significant enrichment of miR-143 seed sites in their 3' UTRs. Here we report the identification of Hexokinase 2 (HK2) as a direct target of miR-143. We show that re-introduction of miR-143 in the colon cancer cell line DLD-1 results in a decreased lactate secretion. CONCLUSION: We have identified...... and validated HK2 as a miR-143 target. Furthermore, our results indicate that miR-143 mediated down-regulation of HK2 affects glucose metabolism in colon cancer cells. We hypothesize that loss of miR-143-mediated repression of HK2 can promote glucose metabolism in cancer cells, contributing to the shift towards......ABSTRACT: BACKGROUND: MicroRNAs (miRNAs) are well recognized as gene regulators and have been implicated in the regulation of development as well as human diseases. miR-143 is located at a fragile site on chromosome 5 frequently deleted in cancer, and has been reported to be down...

  2. Nutlin-3 down-regulates retinoblastoma protein expression and inhibits muscle cell differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Walsh, Erica M. [Department of Biochemistry, Boston University School of Medicine, Boston, MA 02118 (United States); Niu, MengMeng; Bergholz, Johann [Center of Growth, Metabolism and Aging, College of Life Sciences, Sichuan University, Chengdu, 610014 China (China); Jim Xiao, Zhi-Xiong, E-mail: jxiao@bu.edu [Department of Biochemistry, Boston University School of Medicine, Boston, MA 02118 (United States); Center of Growth, Metabolism and Aging, College of Life Sciences, Sichuan University, Chengdu, 610014 China (China)

    2015-05-29

    The p53 tumor suppressor gene plays a critical role in regulation of proliferation, cell death and differentiation. The MDM2 oncoprotein is a major negative regulator for p53 by binding to and targeting p53 for proteasome-mediated degradation. The small molecule inhibitor, nutlin-3, disrupts MDM2-p53 interaction resulting in stabilization and activation of p53 protein. We have previously shown that nutlin-3 activates p53, leading to MDM2 accumulation as concomitant of reduced retinoblastoma (Rb) protein stability. It is well known that Rb is important in muscle development and myoblast differentiation and that rhabdomyosarcoma (RMS), or cancer of the skeletal muscle, typically harbors MDM2 amplification. In this study, we show that nutlin-3 inhibited myoblast proliferation and effectively prevented myoblast differentiation, as evidenced by lack of expression of muscle differentiation markers including myogenin and myosin heavy chain (MyHC), as well as a failure to form multinucleated myotubes, which were associated with dramatic increases in MDM2 expression and decrease in Rb protein levels. These results indicate that nutlin-3 can effectively inhibit muscle cell differentiation. - Highlights: • Nutlin-3 inhibits myoblast proliferation and prevents differentiation into myotubes. • Nutlin-3 increases MDM2 expression and down-regulates Rb protein levels. • This study has implication in nutlin-3 treatment of rhabdomyosarcomas.

  3. Fucoidan induces apoptosis of HepG2 cells by down-regulating p-Stat3.

    Science.gov (United States)

    Roshan, Sadia; Liu, Yun-yi; Banafa, Amal; Chen, Hui-jie; Li, Ke-xiu; Yang, Guang-xiao; He, Guang-yuan; Chen, Ming-jie

    2014-06-01

    Fucoidan is one of the main bioactive components of polysaccharides. The current study was focused on the anti-tumor effects of fucoidan on human heptoma cell line HepG2 and the possible mechanisms. Fucoidan treatment resulted in cell cycle arrest and apoptosis of HepG2 cells in a dose-dependent manner detected by MTT assay, flow cytometry and fluorescent microscopy. The results of flow cytometric analysis revealed that fucoidan induced G2/M arrest in the cell cycle progression. Hoechst 33258 and Annexin V/PI staining results showed that the apoptotic cell number was increased, which was associated with a dose-dependent up-regulation of Bax and down-regulation of Bcl-2 and p-Stat3. In parallel, the up-regulation of p53 and the increase in reactive oxygen species were also observed, which may play important roles in the inhibition of HepG2 growth by fucoidan. In the meantime, Cyclin B1 and CDK1 were down-regulated by fucoidan treatment. Down-regulation of p-Stat3 by fucoidan resulted in apoptosis and an increase in ROS in response to fucoidan exposure. We therefore concluded that fucoidan induces apoptosis through the down-regulation of p-Stat3. These results suggest that fucoidan may be used as a novel anti-cancer agent for hepatocarcinoma.

  4. Salinomycin enhances cisplatin-induced cytotoxicity in human lung cancer cells via down-regulation of AKT-dependent thymidylate synthase expression.

    Science.gov (United States)

    Ko, Jen-Chung; Zheng, Hao-Yu; Chen, Wen-Ching; Peng, Yi-Shuan; Wu, Chia-Hung; Wei, Chia-Li; Chen, Jyh-Cheng; Lin, Yun-Wei

    2016-12-15

    Salinomycin, a polyether antibiotic, acts as a highly selective potassium ionophore and has anticancer activity on various cancer cell lines. Cisplatin has been proved as chemotherapy drug for advanced human non-small cell lung cancer (NSCLC). Thymidylate synthase (TS) is a key enzyme in the pyrimidine salvage pathway, and increased expression of TS is thought to be associated with resistance to cisplatin. In this study, we showed that salinomycin (0.5-2μg/mL) treatment down-regulating of TS expression in an AKT inactivation manner in two NSCLC cell lines, human lung adenocarcinoma A549 and squamous cell carcinoma H1703 cells. Knockdown of TS using small interfering RNA (siRNA) or inhibiting AKT activity with PI3K inhibitor LY294002 enhanced the cytotoxicity and cell growth inhibition of salinomycin. A combination of cisplatin and salinomycin resulted in synergistic enhancement of cytotoxicity and cell growth inhibition in NSCLC cells, accompanied with reduced activation of phospho-AKT, and TS expression. Overexpression of a constitutive active AKT (AKT-CA) expression vector reversed the salinomycin and cisplatin-induced synergistic cytotoxicity. In contrast, pretreatment with LY294002 further decreased the cell viability in salinomycin and cisplatin cotreated cells. Our findings suggested that the down-regulation of AKT-mediated TS expression by salinomycin enhanced the cisplatin-induced cytotoxicity in NSCLC cells. These results may provide a rationale to combine salinomycin with cisplatin for lung cancer treatment.

  5. VCP/p97, down-regulated by microRNA-129-5p, could regulate the progression of hepatocellular carcinoma.

    Directory of Open Access Journals (Sweden)

    Yu Liu

    Full Text Available Valosin containing protein (VCP/p97 plays various important roles in cells. Moreover, elevated expression of VCP in hepatocellular carcinoma (HCC is correlated with increased incidence of recurrence. But the role of VCP in HCC progression in vitro and in vivo is unclear. And there are few reports about the regulation mechanism on the expression of VCP in HCC. In this study, it was identified that the level of VCP was frequently increased in human HCC tissues. In addition, down-regulation of VCP with siRNAs could dramatically suppress the genesis and progression of tumor in vivo. It was found that miR-129-5p directly inhibited the expression of VCP in several HCC cell lines. Meanwhile, the level of VCP in HCC tissues was negatively associated with the level of miR-129-5p. Our further investigation showed that the enhanced expression of miR-129-5p also suppressed tumor growth in vivo. Moreover, it was revealed that miR-129-5p could inhibit the degradation of IκBα and increase the apoptosis and reduce the migration of HCC cells by suppressing the expression of VCP. Our results revealed that the expression of VCP was directly regulated by miR-129-5p and this regulation played an important role in the progression of HCC.

  6. Association of Notch pathway down-regulation with Triple Negative/Basal-like breast carcinomas and high tumor-infiltrating FOXP3+ Tregs.

    Science.gov (United States)

    Ortiz-Martínez, Fernando; Gutiérrez-Aviñó, Francisco José; Sanmartín, Elena; Pomares-Navarro, Eloy; Villalba-Riquelme, Cristina; García-Martínez, Araceli; Lerma, Enrique; Peiró, Gloria

    2016-06-01

    T regulatory cells (Tregs) are a lineage of lymphocytes involved in immune response suppression that are characterized by the expression of the forkhead box P3 (FOXP3) transcription factor. Notch pathway regulates FOXP3 transcription in Tregs, but its role in breast cancer is unknown. We aimed at studying whether Notch pathway regulates FOXP3 expression and Tregs content in breast cancer, and its association with luminal breast carcinomas. We analyzed by quantitative Real-Time PCR the mRNA levels of FOXP3, Notch pathway genes (Notch1, Notch2, Notch4 and Jagged1) and STAT3 in a series of 152 breast carcinomas including hormone receptor-positive and -negative phenotypes (luminal and Triple Negative/Basal-like). We also studied the protein expression of Notch1, STAT3 and FOXP3 by immunohistochemistry. High FOXP3 mRNA levels correlated with larger tumor size (p=0.010), histological grade 3 (p=0.008) and positive lymph-node status (p=0.031). Also, low levels of Notch pathway genes mRNA correlated with poor prognostic factors such as larger tumor size, positive lymph-node status, tumor phenotype and infiltrating tumor Tregs. A survival analysis for the patients showed that large tumor size, histological grade 3, vascular invasion, infiltrating Tregs and low Notch1 mRNA expression were significantly associated with a decreased patients' overall survival (p≤0.05). On a multivariate analysis, high Tregs content (HR=3.00, 95% CI 1.04-8.90, p=0.042) and low Notch1 mRNA levels (HR=3.33, 95% CI 1.02-10.86, p=0.046) were independent markers for overall survival. Our results support that the Notch pathway up-regulation promotes luminal breast carcinomas, whereas down-regulation correlates with the expression of FOXP3, favors tumor Tregs infiltration and associates with Triple Negative/Basal-like tumors.

  7. Endothelial progenitor cell down-regulation in a mouse model of Kawasaki disease

    Institute of Scientific and Technical Information of China (English)

    LIU Jun-feng; DU Zhong-dong; CHEN Zhi; LU Dun-xiang; LI Li; GUAN Yun-qian; WAN Sui-gui

    2012-01-01

    Background Cardiovascular complications of Kawasaki disease (KD) are a common cause of heart disease in pediatric populations.Previous studies have suggested a role for endothelial progenitor cells (EPCs) in coronary artery lesions associated with KD.However,long-term observations of EPCs during the natural progression of this disorder are lacking.Using an experimental model of KD,we aimed to determine whether the coronary artery lesions are associated with down-regulation of EPCs.Methods To induce KD,C57BL/6 mice were administered an intraperitoneal injection of Lactobacillus casei cell wall extract (LCWE; phosphate buffered saline used as control vehicle).Study groups included:group A (14 days following LCWE injection),group B (56 days following LCWE injection) and group C (controls).Numbers of circulating EPCs (positively staining for both CD34 and FIk-1 while staining negative for CD45) were evaluated using flow cytometry.Bone marrow mononuclear cells were cultured in vitro to expand EPCs for functional analysis.In vitro EPC proliferation,adhesion and migration were assessed.Results The model was shown to exhibit similar coronary artery lesions to KD patients with coronary aneurysms.Numbers of circulating EPCs decreased significantly in the KD models (groups A and B) compared to controls ((0.017±0.008)% VS.(0.028±0.007)%,P<0.05 and (0.016±0.007)% vs.(0.028±0.007)%,P <0.05).Proliferative,adhesive and migratory properties of EPCs were markedly impaired in groups A and B.Conclusion Coronary artery lesions in KD occur as a consequence of impaired vascular injury repair,resulting from excess consumption of EPCs together with a functional impairment of bone marrow EPCs and their precursors.

  8. Down-regulated βIII-tubulin Expression Can Reverse Paclitaxel Resistance in A549/Taxol Cells Lines

    Directory of Open Access Journals (Sweden)

    Yinling ZHUO

    2014-08-01

    Full Text Available Background and objective Chemotherapy drug resistance is the primary causes of death in patients with pulmonary carcinoma which make tumor recurrence or metastasis. β-tubulin is the main cell targets of anti-microtubule drug. Increased expression of βIII-tubulin has been implicated in non-small cell lung cancer (NSCLC cell lines. To explore the relationship among the expression level of βIII-tubulin and the sensitivity of A549/Taxolcell lines to Taxol and cell cycles and cell apoptosis by RNA interference-mediated inhibition of βIII-tubulin in A549/Taxol cells. Methods Three pairs of siRNA targetd βIII-tubulin were designed and prepared, which were transfected into A549/Taxol cells using LipofectamineTM 2000. We detected the expression of βIII-tubulin mRNA using Real-time fluorescence qRT-PCR. Tedhen we selected the most efficient siRNA by the expression of βIII-tubulin mRNA in transfected group. βIII-tubulin protein level were mesured by Western blot. The taxol sensitivity in transfected group were evaluated by MTT assay. And the cell apoptosis and cell cycles were determined by flow cytometry. Results βIII-tubulin mRNA levels in A549/Taxol cells were significantly decreased in transfected grop by Real-time qRT-PCR than control groups. And βIII-tubulin siRNA-1 sequence showed the highest transfection efficiency, which was (87.73±4.87% (P<0.01; Western blot results showed that the expressional level of BIII tublin protein was significantly down-reulated in the transfectant cells than thant in the control cells. By MTT assay, we showed that the inhibition ratio of Taxol to A549/Taxol cells transfeced was higher than that of control group (51.77±4.60% (P<0.01. The early apoptosis rate of A549/Taxol cells in transfected group were significantly higher than that of control group (P<0.01; G2-M content in taxol group obviously increased than untreated samples by the cell cycle (P<0.05. Conclusion βIII-tubulin down-regulated significantly

  9. Down-regulation of HSP27 sensitizes TRAIL-resistant tumor cell to TRAIL-induced apoptosis

    DEFF Research Database (Denmark)

    Zhuang, Hongqin; Jiang, Weiwei; Cheng, Wei

    2010-01-01

    oxygen species or anticancer drugs. Their elevated expressions facilitate cells to survive in stress circumstances. The HSP27 expression is enhanced in many tumor cells, implying that it is involved in tumor progression and the development of treatment resistance in various tumors, including lung cancer...... siRNA on drug sensitization of A549 cells to TRAIL treatment. The results showed that treatment of A549 cells with HSP27 siRNA down-regulated HSP27 expression but did not induce significant apoptosis. However, combination of HSP27 siRNA with TRAIL-induced significant apoptosis in TRAIL-resistant A549...... cells. In addition to inducing caspases activation and apoptosis, combined treatment with HSP27 siRNA and TRAIL also increased JNK and p53 expression and activity. Collectively, these findings provide a conclusion that siRNA targeting of the HSP27 gene specifically down-regulated HSP27 expression in A...

  10. Sodium arsenite down-regulates the expression of X-linked inhibitor of apoptosis protein via translational and post-translational mechanisms in hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Hong; Hao, Yuqing; Wang, Lijing; Jia, Dongwei [Gene Research Center, Shanghai Medical College, Fudan University, Shanghai 200032 (China); Ruan, Yuanyuan, E-mail: yuanyuanruan@fudan.edu.cn [Gene Research Center, Shanghai Medical College, Fudan University, Shanghai 200032 (China); Gu, Jianxin [Gene Research Center, Shanghai Medical College, Fudan University, Shanghai 200032 (China)

    2012-06-15

    Highlights: Black-Right-Pointing-Pointer Sodium arsenite down-regulates the protein expression level of XIAP in HCC. Black-Right-Pointing-Pointer Sodium arsenite inhibits the de novo XIAP synthesis and its IRES activity. Black-Right-Pointing-Pointer Sodium arsenite decreases XIAP stability and promotes its proteasomal degradation. Black-Right-Pointing-Pointer Overexpression of XIAP attenuates the pro-apoptotic effect of sodium arsenite. -- Abstract: X-linked inhibitor of apoptosis protein (XIAP) is a member of the inhibitors of apoptosis protein (IAP) family, and has been reported to exhibit elevated expression levels in hepatocellular carcinoma (HCC) and promote cell survival, metastasis and tumor recurrence. Targeting XIAP has proven effective for the inhibition of cancer cell proliferation and restoration of cancer cell chemosensitivity. Arsenic (or sodium arsenite) is a potent anti-tumor agent used to treat patients with acute promyelocytic leukemia (APL). Additionally, arsenic induces cell growth inhibition, cell cycle arrest and apoptosis in human HCC cells. In this study, we identified XIAP as a target for sodium arsenite-induced cytotoxicity in HCC. The exposure of HCC cell lines to sodium arsenite resulted in inhibition of XIAP expression in both a dose- and time-dependent manner. Sodium arsenite blocked the de novo XIAP synthesis and the activity of its internal ribosome entry site (IRES) element. Moreover, treatment with sodium arsenite decreased the protein stability of XIAP and induced its ubiquitin-proteasomal degradation. Overexpression of XIAP attenuated the pro-apoptotic effect of sodium arsenite in HCC. Taken together, our data demonstrate that sodium arsenite suppresses XIAP expression via translational and post-translational mechanisms in HCC.

  11. Nucleolin down-regulation is involved in ADP-induced cell cycle arrest in S phase and cell apoptosis in vascular endothelial cells.

    Directory of Open Access Journals (Sweden)

    Wenmeng Wang

    Full Text Available High concentration of extracellular ADP has been reported to induce cell apoptosis, but the molecular mechanisms remain not fully elucidated. In this study, we found by serendipity that ADP treatment of human umbilical vein endothelial cells (HUVEC and human aortic endothelial cells (HAEC down-regulated the protein level of nucleolin in a dose- and time-dependent manner. ADP treatment did not decrease the transcript level of nucloelin, suggesting that ADP might induce nucleolin protein degradation. HUVEC and HAEC expressed ADP receptor P2Y13 receptor, but did not express P2Y1 or P2Y12 receptors. However, P2Y1, 12, 13 receptor antagonists MRS2179, PSB0739, MRS2211 did not inhibit ADP-induced down-regulation of nucleolin. Moreover, MRS2211 itself down-regulated nucleolin protein level. In addition, 2-MeSADP, an agonist for P2Y1, 12 and 13 receptors, did not down-regulate nucleolin protein. These results suggested that ADP-induced nucleolin down-regulation was not due to the activation of P2Y1, 12, or 13 receptors. We also found that ADP treatment induced cell cycle arrest in S phase, cell apoptosis and cell proliferation inhibition via nucleolin down-regulation. The over-expression of nucleolin by gene transfer partly reversed ADP-induced cell cycle arrest, cell apoptosis and cell proliferation inhibition. Furthermore, ADP sensitized HUVEC to cisplatin-induced cell death by the down-regulation of Bcl-2 expression. Taken together, we found, for the first time to our knowledge, a novel mechanism by which ADP regulates cell proliferation by induction of cell cycle arrest and cell apoptosis via targeting nucelolin.

  12. Lycium barbarum L. Polysaccharide (LBP Reduces Glucose Uptake via Down-Regulation of SGLT-1 in Caco2 Cell

    Directory of Open Access Journals (Sweden)

    Huizhen Cai

    2017-02-01

    Full Text Available Lycium barbarum L. polysaccharide (LBP is prepared from Lycium barbarum L. (L. barbarum, which is a traditional Chinese medicine. LPB has been shown to have hypoglycemic effects. In order to gain some mechanistic insights on the hypoglycemic effects of LBP, we investigated the uptake of LBP and its effect on glucose absorption in the human intestinal epithelial cell line Caco2 cell. The uptake of LBP through Caco2 cell monolayer was time-dependent and was inhibited by phloridzin, a competitive inhibitor of SGLT-1. LPB decreased the absorption of glucose in Caco2 cell, and down-regulated the expression of SGLT-1. These results suggest that LBP might be transported across the human intestinal epithelium through SGLT-1 and it inhibits glucose uptake via down-regulating SGLT-1.

  13. Down-regulation of adipogenesis of mesenchymal stem cells by oscillating high-gradient magnetic fields and mechanical vibration

    Science.gov (United States)

    Zablotskii, V.; Lunov, O.; Novotná, B.; Churpita, O.; Trošan, P.; HoláÅ, V.; Syková, E.; Dejneka, A.; Kubinová, Š.

    2014-09-01

    Nowadays, the focus in medicine on molecular genetics has resulted in a disregard for the physical basis of treatment even though many diseases originate from changes in cellular mechanics. Perturbations of the cellular nanomechanics promote pathologies, including cardiovascular disease and cancer. Furthermore, whilst the biological and therapeutic effects of magnetic fields are a well-established fact, to date the underlying mechanisms remain obscure. Here, we show that oscillating high-gradient magnetic field (HGMF) and mechanical vibration affect adipogenic differentiation of mesenchymal stem cells by the transmission of mechanical stress to the cell cytoskeleton, resulting in F-actin remodelling and subsequent down-regulation of adipogenic genes adiponectin, PPARγ, and AP2. Our findings propose an insight into the regulation of cellular nanomechanics, and provide a basis for better controlled down-regulation of stem cell adipogenesis by HGMF, which may facilitate the development of challenging therapeutic strategies suitable for the remote control of biological systems.

  14. Sensitization of multidrug-resistant human cancer cells to Hsp90 inhibitors by down-regulation of SIRT1.

    Science.gov (United States)

    Kim, Hak-Bong; Lee, Su-Hoon; Um, Jee-Hyun; Oh, Won Keun; Kim, Dong-Wan; Kang, Chi-Dug; Kim, Sun-Hee

    2015-11-03

    The effectiveness of Hsp90 inhibitors as anticancer agents was limited in multidrug-resistant (MDR) human cancer cells due to induction of heat shock proteins (Hsps) such as Hsp70/Hsp27 and P-glycoprotein (P-gp)-mediated efflux. In the present study, we showed that resistance to Hsp90 inhibitors of MDR human cancer cells could be overcome with SIRT1 inhibition. SIRT1 knock-down or SIRT1 inhibitors (amurensin G and EX527) effectively suppressed the resistance to Hsp90 inhibitors (17-AAG and AUY922) in several MDR variants of human lymphoblastic leukemia and human breast cancer cell lines. SIRT1 inhibition down-regulated the expression of heat shock factor 1 (HSF1) and subsequently Hsps and facilitated Hsp90 multichaperone complex disruption via hyperacetylation of Hsp90/Hsp70. These findings were followed by acceleration of ubiquitin ligase CHIP-mediated mutant p53 (mut p53) degradation and subsequent down-regulation of P-gp in 17-AAG-treated MDR cancer cells expressing P-gp and mut p53 after inhibition of SIRT1. Therefore, combined treatment with Hsp90 inhibitor and SIRT1 inhibitor could be a more effective therapeutic approach for Hsp90 inhibitor-resistant MDR cells via down-regulation of HSF1/Hsps, mut p53 and P-gp.

  15. Collagen I-induced dendritic cells activation is regulated by TNF- production through down-regulation of IRF4

    Indian Academy of Sciences (India)

    Barun Poudel; Hyeon-Hui Ki; Young-Mi Lee; Dae-Ki Kim

    2015-03-01

    Previously we have shown that collagen I enhances the maturation and function of dendritic cells (DCs). Inflammatory mediators such as tumour necrosis factor (TNF)-, interleukin (IL)-1 and lipopolysaccharide (LPS) are also known to activate DCs. Here we investigated the involvement of TNF- on the collagen I-induced DCs activation. TNF-a neutralization inhibited collagen I-induced IL-12 secretions by DCs. Additionally, we observed suppression of collagen I-induced costimulatory molecules expression along with down-regulation of genes involved in DCs activation pathway. Furthermore, TNF- inhibition upon collagen Istimulation up-regulated the expression of interferon regulatory transcription factor IRF4, when compared to collagen I only treated cells. Collectively, our data demonstrate that collagen I induce TNF- production, which is crucial for the activation and function of DCs, through down-regulation of IRF4, and implicates the importance in development of anti- TNF- therapeutics for several inflammatory diseases.

  16. MiR-378 Promotes the Migration of Liver Cancer Cells by Down-Regulating Fus Expression

    Directory of Open Access Journals (Sweden)

    Jichun Ma

    2014-12-01

    Full Text Available Background: miR-378 regulates osteoblast differentiation and participates in tumor cell self-renewal and chemo-resistance. However, the function of miR-378 in liver cancer cell migration has not been reported to date. Methods: miR-378 expression was examined using real-time quantitative PCR. HepG2 cell migration and liver cell invasion were examined using wound-healing and cell invasion assays. Additionally, HepG2 cell metastasis was analyzed in nude mice. Results: miR-378 over-expression enhances HepG2 cell proliferation, migration and liver cell invasion. Typical metastatic lesions were found in the livers of mice injected with miR-378-transfected cells, and high levels of the CMV promoter were detected in the nodules, indicating that miR-378 promoted the metastasis of the tumor cells to the liver. We also demonstrated that miR-378 down-regulated Fus expression. Conclusions: Our results suggested that miR-378 enhanced cell migration and metastasis by down-regulating Fus expression.

  17. Down-Regulation of Notchl and NF-κB by Curcumin in Breast Cancer Cells MDA-MB-231

    Institute of Scientific and Technical Information of China (English)

    LONG Li; CAO You-de

    2008-01-01

    Objective:To test whether the down-regulation of Notch1 gene expression by curcumin could inhibit cell growth and induce apoptosis,which may be associated mechanistically with the down-regulation of NF-κB in breast cancer cells. Methods:Breast cancer cell lines MDA-MB-231 were cultured in vitro and treated with different dosages of curcumin(0,1.25,5.0,20.0μmol/L)for dose-dependent assay and different time(0,24,48,72 h)at the dosage of 5.0μmol/L for time course assay.The changes of the mRNA and protein expression of Notch1 and NF-κB were measured by RT-PCR and Western Blot,and MTT assay was used to measure the change of proliferation. Results:The mRNA and protein levels of Notch 1 and NF-κB were decreased significantly in human breast cancer cell line with the increase of dosage of curcumin(P<0.05),and with the extension of time course(P<0.05).These changes suggested a dose- and time-dependent manner.The proliferation rate of cells also was significantly inhibited(P<0.05). Conclusion:The current results show that the Notch-1 signaling pathway is associated mechanistically with NF-κB activity during curcumin-induced cell growth inhibition and apoptosis of breast cancer cells.These results suggest that the down-regulation of Notch signaling by curcumin may be a novel strategy for the treatment of patients with breast cancer.

  18. Down regulation of miR-203 in radiation-induced thymic lymphoma promoted cells proliferation and inhibited apoptosis%Down regulation of miR-203in radiation-induced thymic lymphoma promoted cells proliferation and inhibited apoptosis

    Institute of Scientific and Technical Information of China (English)

    Zhang Chaoxiong; Zhang Mingjian; Gao Fu; Zhou Chuanfeng; Zhang Pei; Cai Jianming; Liu Cong

    2015-01-01

    Objective To investigate the role of miR-203 in radiation-induced thymic lymphoma (RITL).Methods A 60Co irradiator was used for total-body irradiation.MicroRNAs(miRNAs) level was assayed by qRT-PCR.Cell proliferation was assayed by MTT assay.Cell apoptosis was examined by fluorescence activated cell sorter (FACS).Dual luciferase reporter assay system was used to detect the 3'UTR reporter.Results MiR-203 was down-regulated in RITL tissues.Overexpression of miR-203 strongly inhibited the proliferation of both NIH3T3 cells and EL4 cells and vice versa.MiR-203 inhibited cells proliferation and induced apoptosis via TANK-binding kinase (TBK1),SLUG (SNAI2) and Cyclin D1 (CCND1).Conclusions Radiation down-regulated the level of miR-203 in thymic,which promoted radiation-induced thymic lymphoma by targeting TBK1,SNAI2 and CCND1.

  19. N-methylhemeanthidine chloride, a novel Amaryllidaceae alkaloid, inhibits pancreatic cancer cell proliferation via down-regulating AKT activation

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Guoli; Yao, Guangmin; Zhan, Guanqun; Hu, Yufeng [Hubei Key Laboratory of Natural Medicinal Chemistry and Resource Evaluation, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei PR China (China); Yue, Ming [Hubei Key Laboratory of Drug Target Research and Pharmacodynamic Evaluation, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei (China); Cheng, Ling; Liu, Yaping; Ye, Qi [Hubei Key Laboratory of Natural Medicinal Chemistry and Resource Evaluation, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei PR China (China); Qing, Guoliang [Hubei Key Laboratory of Drug Target Research and Pharmacodynamic Evaluation, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei (China); Zhang, Yonghui, E-mail: zhangyh@mails.tjmu.edu.cn [Hubei Key Laboratory of Natural Medicinal Chemistry and Resource Evaluation, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei PR China (China); Liu, Hudan, E-mail: hudanliu@hust.edu.cn [Hubei Key Laboratory of Natural Medicinal Chemistry and Resource Evaluation, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei PR China (China)

    2014-11-01

    We previously reported the isolation of a novel Amaryllidaceae alkaloid, N-methylhemeanthidine chloride (NMHC), from Zephyranthes candida, which exhibits potent cytotoxicity in a spectrum of tumor cells. However, the mechanism of action remains unclear. Using multiple cell lines derived from human pancreatic cancer, one of the most mortal and refractory human malignancies, we further studied the NMHC-mediated cytotoxicity and found that it induced drastic cytotoxicity in pancreatic cancer cells whereas an insignificant effect on a noncancerous cell line. The NMHC-mediated growth inhibition was more severe than the first-line chemotherapeutic agent gemcitabine, leading to cell cycle arrest, apoptotic death and decreased glycolysis. NMHC exerted its function through down-regulating AKT activation, and the ectopic expression of activated AKT rescued the growth inhibition. Consistently, NMHC injections in a pancreatic cancer xenograft model manifested the anti-tumor effect in vivo. Engrafted tumor cells underwent AKT attenuation and apoptotic death upon treatments. As such, we here demonstrate the AKT inhibition may be one of the mechanisms by which NMHC decreases tumor cell survival rate in vitro and in vivo. Our data thereby suggest that NMHC holds great promise as a potent chemotherapeutic agent against pancreatic cancer and sheds new light on obtaining such agents from natural products toward therapeutic purposes. - Highlights: • N-methylhemeanthidine chloride (NMHC) is a novel Amaryllidaceae alkaloid. • NMHC exhibits potent anti-neoplastic activity. • NMHC leads to cell cycle arrest, apoptotic death and decreased metabolism. • NMHC down-regulates the AKT signaling pathway.

  20. Down-regulation of Sonic hedgehog signaling pathway activity is involved in 5-fluorouracil-induced apoptosis and motility inhibition in Hep3B cells

    Institute of Scientific and Technical Information of China (English)

    Qiyu Wang; Shuhong Huang; Ling Yang; Ling Zhao; Yuxia Yin; Zhongzhen Liu; Zheyu Chen; Hongwei Zhang

    2008-01-01

    The Sonic hedgehog (SHh) pathway plays a critical role in normal embryogenesis and carcinogenesis, but its function in cancer cells treated with 5-fluorouracil (5-FU) remains unknown. We examined the expression of a subset of SHh signaling pathway genes, including SHh, SMO, PTC1, Su(Fu) and HIP in human hepatocellular carcinoma (HCC) cell lines,Hep3B and HepG2, treated with 5-FU by reverse transcriptionpolymerase chain reaction. Using trypan blue analysis,3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling assay, we also detected the apoptosis of Hep3B cells resulting from the transfection of pCS2-Gli1 expression vector combined with 5-FU treatment.The motility of the cells was detected by scratch wound closure assay. The expression and subcellular location of PTC1 protein in Hep3B cells treated by 5-FU were also investigated by Western blot analysis and immunofluorescent microscopy. The results indicated that the expression of SHh pathway target molecules at both messenger RNA and protein levels are evidently down-regulated in Hep3B cells treated with 5-FU. The overexpression of Gli1 restores cell viability and, to some extent, the migration abilities inhibited by 5-FU.Furthermore, 5-FU treatment affects the subcellular localization of PTC1 protein, a key member in SHh signaling pathway. Our data showed that the down-regulation of SHh signaling pathway activity was involved in 5-FU-induced apoptosis and the inhibition of motility in hedgehog-activated HCC cell lines. This implies that the combination of SHh signaling pathway inhibitor and 5-FU-based chemotherapy might represent a more promising strategy against HCC.

  1. Curcumin inhibits cell growth and invasion and induces apoptosis through down-regulation of Skp2 in pancreatic cancer cells

    Science.gov (United States)

    Su, Jingna; Zhou, Xiuxia; Wang, Lixia; Yin, Xuyuan; Wang, Zhiwei

    2016-01-01

    Natural polyphenol compound curcumin has been found to exhibit its anticancer activity in a variety of human malignancies including pancreatic cancer (PC). However, the underlying mechanism has not been fully understood. Accumulating evidence has demonstrated that Skp2 (S-phase kinase associated protein 2) plays an oncogenic role in the development and progression of human cancers. In this study, we aim to explore the molecular basis of curcumin-induced cell growth inhibition in PC cells.Multiple methods such as CTG assay, Flow cytometry, clonogenic assay, wound healing assay, Transwell invasion assay, Western blotting, and transfection were performed to validate the oncogenic role of curcumin in PC cells. We found that curcumin suppressed cell growth, clonogenic potential, migration and invasion, and induced cell apoptosis and cell cycle arrest. Moreover, we observed thatover-expression of Skp2 significantly promoted cell growth, whereas down-regulation of Skp2 with siRNAs inhibited cell growth. The molecular basis of curcumin-mediated cell growth inhibition we identified is that curcumin significantly suppressed Skp2 expression and subsequently induced p21 expression. These findings suggested thattargeting Skp2 by curcumin could be a promising therapeutic strategy for the treatment of PC patients.

  2. Endotoxin-induced basal respiration alterations of renal HK-2 cells: A sign of pathologic metabolism down-regulation

    Energy Technology Data Exchange (ETDEWEB)

    Quoilin, C., E-mail: cquoilin@ulg.ac.be [Laboratory of Biomedical Spectroscopy, Department of Physics, University of Liege, 4000 Liege (Belgium); Mouithys-Mickalad, A. [Center of Oxygen Research and Development, Department of Chemistry, University of Liege, 4000 Liege (Belgium); Duranteau, J. [Department of Anaesthesia and Surgical ICU, CHU Bicetre, University Paris XI Sud, 94275 Le Kremlin Bicetre (France); Gallez, B. [Biomedical Magnetic Resonance Group, Louvain Drug Research Institute, Universite catholique de Louvain, 1200 Brussels (Belgium); Hoebeke, M. [Laboratory of Biomedical Spectroscopy, Department of Physics, University of Liege, 4000 Liege (Belgium)

    2012-06-29

    Highlights: Black-Right-Pointing-Pointer A HK-2 cells model of inflammation-induced acute kidney injury. Black-Right-Pointing-Pointer Two oximetry methods: high resolution respirometry and ESR spectroscopy. Black-Right-Pointing-Pointer Oxygen consumption rates of renal cells decrease when treated with LPS. Black-Right-Pointing-Pointer Cells do not recover normal respiration when the LPS treatment is removed. Black-Right-Pointing-Pointer This basal respiration alteration is a sign of pathologic metabolism down-regulation. -- Abstract: To study the mechanism of oxygen regulation in inflammation-induced acute kidney injury, we investigate the effects of a bacterial endotoxin (lipopolysaccharide, LPS) on the basal respiration of proximal tubular epithelial cells (HK-2) both by high-resolution respirometry and electron spin resonance spectroscopy. These two complementary methods have shown that HK-2 cells exhibit a decreased oxygen consumption rate when treated with LPS. Surprisingly, this cellular respiration alteration persists even after the stress factor was removed. We suggested that this irreversible decrease in renal oxygen consumption after LPS challenge is related to a pathologic metabolic down-regulation such as a lack of oxygen utilization by cells.

  3. Histone Deacetylase (HDAC Inhibitors Down-Regulate Endothelial Lineage Commitment of Umbilical Cord Blood Derived Endothelial Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Horia Maniu

    2012-11-01

    Full Text Available To test the involvement of histone deacetylases (HDACs activity in endothelial lineage progression, we investigated the effects of HDAC inhibitors on endothelial progenitors cells (EPCs derived from umbilical cord blood (UCB. Adherent EPCs, that expressed the endothelial marker proteins (PCAM-1, CD105, CD133, and VEGFR2 revealed by flow cytometry were treated with three HDAC inhibitors: Butyrate (BuA, Trichostatin A (TSA, and Valproic acid (VPA. RT-PCR assay showed that HDAC inhibitors down-regulated the expression of endothelial genes such as VE-cadherin, CD133, CXCR4 and Tie-2. Furthermore, flow cytometry analysis illustrated that HDAC inhibitors selectively reduce the expression of VEGFR2, CD117, VE-cadherin, and ICAM-1, whereas the expression of CD34 and CD45 remained unchanged, demonstrating that HDAC is involved in endothelial differentiation of progenitor cells. Real-Time PCR demonstrated that TSA down-regulated telomerase activity probably via suppression of hTERT expression, suggesting that HDAC inhibitor decreased cell proliferation. Cell motility was also decreased after treatment with HDAC inhibitors as shown by wound-healing assay. The balance of acethylation/deacethylation kept in control by the activity of HAT (histone acetyltransferases/HDAC enzymes play an important role in differentiation of stem cells by regulating proliferation and endothelial lineage commitment.

  4. Down-regulated expression of NPM1 in IMS-M2 cell line by (-)-epigallocatechin-3-gallate

    Institute of Scientific and Technical Information of China (English)

    Hoang Thanh Chi; Bui Thi Kim Ly; Hoang Anh Vu; Yuko Sato; Phu Chi Dung; Phan Thi Xinh

    2014-01-01

    Objective: To investigate the inhibited effect of epigallocatechin-3-gallate (EGCG) on the expression of NPM1 in IMS-M2 cells harboring the NPM1 mutations.Methods:Cell proliferation assay was performed to test the effects of EGCG on cell growth of IMS-M2 cells harboring the NPM1 mutations. Western blot analysis were performed to test the protein expression of NPM1, AKT, those associated with apoptosis. Results: EGCG can down-regulate the expression of NPM1 in IMS-M2 cells harboring the NPM1 mutations. Moreover, EGCG also suppressed the cell proliferation and induced apoptosis in IMS-M2 cells.Conclusions:The results suggested that EGCG could be considered as a reagent for treatment of AML patients with NPM1 mutations.

  5. Down-regulation of PPARgamma1 suppresses cell growth and induces apoptosis in MCF-7 breast cancer cells

    Directory of Open Access Journals (Sweden)

    Wallis Natalie K

    2008-12-01

    Full Text Available Abstract Background Peroxisome proliferator-activated receptor gamma (PPARγ is a member of the nuclear hormone receptor superfamily and is highly expressed in many human tumors including breast cancer. PPARγ has been identified as a potential target for breast cancer therapy based on the fact that its activation by synthetic ligands affects the differentiation, proliferation, and apoptosis of cancer cells. However, the controversial nature of current studies and disappointing results from clinical trials raise questions about the contribution of PPARγ signaling in breast cancer development in the absence of stimulation by exogenous ligands. Recent reports from both in vitro and in vivo studies are inconsistent and suggest that endogenous activation of PPARγ plays a much more complex role in initiation and progression of cancer than previously thought. Results We have previously demonstrated that an increase in expression of PPARγ1 in MCF-7 breast cancer cells is driven by a tumor-specific promoter. Myc-associated zinc finger protein (MAZ was identified as a transcriptional mediator of PPARγ1 expression in these cells. In this study, using RNA interference (RNAi to inhibit PPARγ1 expression directly or via down-regulation of MAZ, we report for the first time that a decrease in PPARγ1 expression results in reduced cellular proliferation in MCF-7 breast cancer cells. Furthermore, we demonstrate that these changes in proliferation are associated with a significant decrease in cell transition from G1 to the S phase. Using a dominant-negative mutant of PPARγ1, Δ462, we confirmed that PPARγ1 acts as a pro-survival factor and showed that this phenomenon is not limited to MCF-7 cells. Finally, we demonstrate that down-regulation of PPARγ1 expression leads to an induction of apoptosis in MCF-7 cells, confirmed by analyzing Bcl-2 expression and PARP-1 cleavage. Conclusion Thus, these findings suggest that an increase in PPARγ1 signaling

  6. Down-regulation of the A3 adenosine receptor in human mast cells upregulates mediators of angiogenesis and remodeling.

    Science.gov (United States)

    Rudich, Noam; Dekel, Ornit; Sagi-Eisenberg, Ronit

    2015-05-01

    Adenosine activated mast cells have been long implicated in allergic asthma and studies in rodent mast cells have assigned the A3 adenosine receptor (A3R) a primary role in mediating adenosine responses. Here we analyzed the functional impact of A3R activation on genes that are implicated in tissue remodeling in severe asthma in the human mast cell line HMC-1 that shares similarities with lung derived human mast cells. Quantitative real time PCR demonstrated upregulation of IL6, IL8, VEGF, amphiregulin and osteopontin. Moreover, further upregulation of these genes was noted upon the addition of dexamethasone. Unexpectedly, activated A3R down regulated its own expression and knockdown of the receptor replicated the pattern of agonist induced gene upregulation. This study therefore identifies the human mast cell A3R as regulator of tissue remodeling gene expression in human mast cells and demonstrates a heretofore-unrecognized mode of feedback regulation that is exerted by this receptor.

  7. Down-regulation of stearoyl-CoA desaturase 1 expression inhibits cell proliferation and induces apoptosis of esophageal squamous cell carcinoma and their related molecular mechanism%抑制酰基-辅酶A去饱和酶1表达对食管鳞癌细胞增殖和凋亡的影响及其分子机制探讨

    Institute of Scientific and Technical Information of China (English)

    郜辉; 冯应勤; 朱金峰; 何兴端; 鲁培; 刘红涛

    2012-01-01

    Objective: To investigate the expression of stearoyl-CoA desaturase-1 (SCD-1) in esophageal squamous cell carcinoma (ESCC) cells and tissues and the effect of downregulation of SCD-1 expression on cell proliferation and apoptosis of ESCC EC1 cells, and to explore their related molecular mechanisms. Methods: The expressions of SCD-1 mRNA and protein were detected in 60 cases of ESCC tissues and the corresponding paracancerous normal esophageal epithelial tissues by immunohistochemistry and in situ hybridization method, respectively. The expressions of SCD-1 mRNA and protein in normal esophageal epithelium and different esophageal squamous cell carcinoma cell lines were detected by real-time fluorescence quantitative PCR (RFQ-PCR) and Western blotting, respectively. After the ESCC EC1 cells were transfected with different concentrations of SCD-1 siRNAs, the expression levels of SCD-1 mRNA and protein were detected by RFQ-PCR and Western blotting, respectively. Subsequently, CCK-8 kit was utilized to analyze the change of cell proliferation of EC1 cells, and the flow cytometry (FCM) was used to detect the change of cell apoptosis of EC1 cells. Finally, the expressions of total Akt, p-Akt, bcl-2 and bax proteins were examined by Western blotting. Results: The percentages of positive cells expressing SCD-1 mRNA and protein were significantly higher in ESCC tissues than those in normal esophageal epithelial tissues (P < 0.05), and the up-regulation of SCD-1 mRNA and protein expression levels were closely associated with histological grade, TNM stage and lymph node metastasis (P < 0.05). Furthermore, as compared with the normal esophageal epithelium, the expressions of SCD-1 mRNA and protein were both markedly up-regulated in ESCC cell lines EC9706, Eca109 and EC1 (P < 0.05), in which the EC1 cells displayed the highest SCD-1 expression level. The expression levels of SCD-1 mRNA and protein in EC1 cells were obviously down-regulated after tranfection with 50 nmol/L SCD-1

  8. The inhibitory mechanism of down-regulation expression of pp-GalNAc-T14 by oridonin on human renal carcinoma cells in vitro%冬凌草甲素下调N-乙酰氨基半乳糖转移酶14(pp-GalNAc-T14)表达对肾癌细胞的体外抑制作用

    Institute of Scientific and Technical Information of China (English)

    李鑫; 郭剑明; 徐凯; 王国民

    2012-01-01

    Objective To investigate the inhibitory mechanism of down-regulation expression of polypeptide N-acetylgalactosaminyl-transferase 14 ( pp-GaINAc-T14) by oridonin on human renal carcinoma cells in vitro. Methods The expression of pp-GaINAc-T14 mRNA was detected by reverse-transcribed polymerase chain reaction(RT-PCR) in 20 resected renal cancer tissue samples and matched normal paraneoplastic tissues. After treated with different concentrations of oridonin, inhibition of human renal cancer cell line (786-0) growth was examined by MTT assay. Then levels of pp-GaINAc-T14 mRNA before and after oridonin treatment was detected by RT-PCR. Results (1) pp-GaINAc- T 1 4 mRNA was detectable in both renal cell carcinoma tissues and matched normal tissues . Compared with the normal control samples,a 34. 91% increase of pp-GaINAc-T14 expression in the renal cell carcinoma tissues was observed (P<0. 05). (2) At 24 hours,48 hours and 72 hours after oridonin treatment in the 786-0 cells, expression of pp-GalNac-T14 mRNA significantly decreased 15. 98%, 44. 30% and 58.61% respectively, compared with control cells (P<0. 05). (3)The growth and proliferation of human renal cell carcinoma cell line 786-0 could be inhibited by oridonin and showed time- and dose-dependent. Conclusions Oridonin inhibit growth of renal cell carcinoma cells through down-regulating the expression of pp-GaINAc-Tl4 and may be used as an anti-tumor medicine in the chemotherapy of renal cell carcinoma.%目的 探讨冬凌草甲素下调N-乙酰氨基半乳糖转移酶14( polypeptide N-acetylgalactosaminy-transferase 14,pp-GalNAc-T14)表达对人肾癌细胞的体外抑制作用.方法 收集20例根治性肾癌切除标本的肾癌组织和相对应的癌旁正常组织,用RT-PCR法检测肾癌组织和癌旁正常肾组织中pp-GalNAc-T14 mRNA的表达情况;用单四唑(MTT)法观察冬凌草甲素对该细胞系(786-0人肾透明细胞癌细胞系)的抑制杀伤效果,并运用RT-PCR法

  9. Cryptococcus neoformans activates bone marrow-derived conventional dendritic cells rather than plasmacytoid dendritic cells and down-regulates macrophages.

    Science.gov (United States)

    Siegemund, Sabine; Alber, Gottfried

    2008-04-01

    Induction of IL-12 and IL-23 is essential for protective immunity against Cryptococcusneoformans. The contribution of dendritic cells vs. macrophages to IL-12/23 production in response to C. neoformans infection is unclear. Activation of conventional bone marrow-derived dendritic cells (BMDC), plasmacytoid BMDC, and bone marrow-derived macrophages (BMMPhi) was assessed by analyzing cytokine responses and the expression of MHC-II, CD86, and CD80 in each cell type. Cryptococcus neoformans induced the release of IL-12/23p40 by BMDC, but not by BMMPhi, in a TLR2- and TLR4-independent but MyD88-dependent manner. Conventional BMDC rather than plasmacytoid BMDC up-regulated MHC-II and CD86, while BMMPhi down-regulated MHC-II and CD86 in response to C. neoformans. The up-regulation of MHC-II and CD86 on BMDC required MyD88. Our data point to conventional DC as critical IL-12/23-producing antigen-presenting cells during cryptococcosis.

  10. Antioxidants Abrogate Alpha-Tocopherylquinone-Mediated Down-Regulation of the Androgen Receptor in Androgen-Responsive Prostate Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Alexandra M Fajardo

    Full Text Available Tocopherylquinone (TQ, the oxidation product of alpha-tocopherol (AT, is a bioactive molecule with distinct properties from AT. In this study, AT and TQ are investigated for their comparative effects on growth and androgenic activity in prostate cancer cells. TQ potently inhibited the growth of androgen-responsive prostate cancer cell lines (e.g., LAPC4 and LNCaP cells, whereas the growth of androgen-independent prostate cancer cells (e.g., DU145 cells was not affected by TQ. Due to the growth inhibitory effects induced by TQ on androgen-responsive cells, the anti-androgenic properties of TQ were examined. TQ inhibited the androgen-induced activation of an androgen-responsive reporter and inhibited the release of prostate specific antigen from LNCaP cells. TQ pretreatment was also found to inhibit AR activation as measured using the Multifunctional Androgen Receptor Screening assay. Furthermore, TQ decreased androgen-responsive gene expression, including TM4SF1, KLK2, and PSA over 5-fold, whereas AT did not affect the expression of androgen-responsive genes. Of importance, the antiandrogenic effects of TQ on prostate cancer cells were found to result from androgen receptor protein down-regulation produced by TQ that was not observed with AT treatment. Moreover, none of the androgenic endpoints assessed were affected by AT. The down-regulation of androgen receptor protein by TQ was abrogated by co-treatment with antioxidants. Overall, the biological actions of TQ were found to be distinct from AT, where TQ was found to be a potent inhibitor of cell growth and androgenic activity in androgen-responsive prostate cancer cells.

  11. Down-regulation of msrb3 and destruction of normal auditory system development through hair cell apoptosis in zebrafish.

    Science.gov (United States)

    Shen, Xiaofang; Liu, Fei; Wang, Yingzhi; Wang, Huijun; Ma, Jing; Xia, Wenjun; Zhang, Jin; Jiang, Nan; Sun, Shaoyang; Wang, Xu; Ma, Duan

    2015-01-01

    Hearing defects can significantly influence quality of life for those who experience them. At this time, 177 deafness genes have been cloned, including 134 non-syndromic hearing-loss genes. The methionine sulfoxide reductase B3 (Ahmed et al., 2011) gene (also called DFNB74) is one such newly discovered hearing-loss gene. Within this gene c.265 T>G and c.55 T>C mutations are associated with autosomal recessive hearing loss. However, the biological role and mechanism underlying how it contributes to deafness is unclear. Thus, to better understand this mutation, we designed splicing morpholinos for the purpose of down-regulating msrb3 in zebrafish. Morphants exhibited small, tiny, fused, or misplaced otoliths and abnormal numbers of otoliths. Down-regulation of msrb3 also caused shorter, thinner, and more crowded cilia. Furthermore, L1-8 neuromasts were reduced and disordered in the lateral line system; hair cells in each neuromast underwent apoptosis. Co-injection with human MSRB3 mRNA partially rescued auditory system defects, but mutant MSRB3 mRNA could not. Thus, msrb3 is instrumental for auditory system development in zebrafish and MSRB3-related deafness may be caused by promotion of hair cell apoptosis.

  12. Quantitative cell signalling analysis reveals down-regulation of MAPK pathway activation in colorectal cancer.

    LENUS (Irish Health Repository)

    Gulmann, Christian

    2009-08-01

    Mitogen-activated protein kinases (MAPK) are considered to play significant roles in colonic carcinogenesis and kinase inhibitor therapy has been proposed as a potential tool in the treatment of this disease. Reverse-phase microarray assays using phospho-specific antibodies can directly measure levels of phosphorylated protein isoforms. In the current study, samples from 35 cases of untreated colorectal cancer colectomies were laser capture-microdissected to isolate epithelium and stroma from cancer as well as normal (i.e. uninvolved) mucosa. Lysates generated from these four tissue types were spotted onto reverse-phase protein microarrays and probed with a panel of antibodies to ERK, p-ERK, p38, p-p38, p-JNK, MEK and p-MEK. Whereas total protein levels were unchanged, or slightly elevated (p38, p = 0.0025) in cancers, activated isoforms, including p-ERK, p-p38 and p-JNK, were decreased two- to four-fold in cancers compared with uninvolved mucosa (p < 0.0023 in all cases except for p-JNK in epithelium, where decrement was non-significant). This was backed up by western blotting. Dukes\\' stage B and C cancers displayed lower p-ERK and p-p38 expression than Dukes\\' stage A cancers, although this was not statistically significant. It is concluded that MAPK activity may be down-regulated in colorectal cancer and that further exploration of inhibitory therapy in this system should be carefully evaluated if this finding is confirmed in larger series.

  13. Triptolide inhibits the proliferation of prostate cancer cells and down-regulates SUMO-specific protease 1 expression.

    Directory of Open Access Journals (Sweden)

    Weiwei Huang

    Full Text Available Recently, traditional Chinese medicine and medicinal herbs have attracted more attentions worldwide for its anti-tumor efficacy. Celastrol and Triptolide, two active components extracted from the Chinese herb Tripterygium wilfordii Hook F (known as Lei Gong Teng or Thunder of God Vine, have shown anti-tumor effects. Celastrol was identified as a natural 26 s proteasome inhibitor which promotes cell apoptosis and inhibits tumor growth. The effect and mechanism of Triptolide on prostate cancer (PCa is not well studied. Here we demonstrated that Triptolide, more potent than Celastrol, inhibited cell growth and induced cell death in LNCaP and PC-3 cell lines. Triptolide also significantly inhibited the xenografted PC-3 tumor growth in nude mice. Moreover, Triptolide induced PCa cell apoptosis through caspases activation and PARP cleavage. Unbalance between SUMOylation and deSUMOylation was reported to play an important role in PCa progression. SUMO-specific protease 1 (SENP1 was thought to be a potential marker and therapeutical target of PCa. Importantly, we observed that Triptolide down-regulated SENP1 expression in both mRNA and protein levels in dose-dependent and time-dependent manners, resulting in an enhanced cellular SUMOylation in PCa cells. Meanwhile, Triptolide decreased AR and c-Jun expression at similar manners, and suppressed AR and c-Jun transcription activity. Furthermore, knockdown or ectopic SENP1, c-Jun and AR expression in PCa cells inhibited the Triptolide anti-PCa effects. Taken together, our data suggest that Triptolide is a natural compound with potential therapeutic value for PCa. Its anti-tumor activity may be attributed to mechanisms involving down-regulation of SENP1 that restores SUMOylation and deSUMOyaltion balance and negative regulation of AR and c-Jun expression that inhibits the AR and c-Jun mediated transcription in PCa.

  14. Down-regulation of malignant potential by alpha linolenic acid in human and mouse colon cancer cells.

    Science.gov (United States)

    Chamberland, John P; Moon, Hyun-Seuk

    2015-03-01

    Omega-3 fatty acids (also called ω-3 fatty acis or n-3 fatty acid) are polyunsaturated fatty acids (PUFAs) with a double bond (C=C) at the third carbon atom from the end of the carbon chain. Numerous test tube and animal studies have shown that omega-3 fatty acids may prevent or inhibit the growth of cancers, suggesting that omega-3 fatty acids are important in cancer physiology. Alpha-linolenic acid (ALA) is one of an essential omega-3 fatty acid and organic compound found in seeds (chia and flaxseed), nuts (notably walnuts), and many common vegetable oils. ALA has also been shown to down-regulate cell proliferation of prostate, breast, and bladder cancer cells. However, direct evidence that ALA suppresses to the development of colon cancer has not been studied. Also, no previous studies have evaluated whether ALA may regulate malignant potential (adhesion, invasion and colony formation) in colon cancer cells. In order to address the questions above, we conducted in vitro studies and evaluated whether ALA may down-regulate malignant potential in human (HT29 and HCT116) and mouse (MCA38) colon cancer cell lines. We observed that treatment with 1-5 mM of ALA inhibits cell proliferation, adhesion and invasion in both human and mouse colon cancer cell lines. Interestingly, we observed that ALA did not decrease total colony numbers when compared to control. By contrast, we found that size of colony was significantly changed by ALA treatment when compared to control in all colon cancer cell lines. We suggest that our data enhance our current knowledge of ALA's mechanism and provide crucial information to further the development of new therapies for the management or chemoprevention of colon cancer.

  15. DMBT1 expression is down-regulated in breast cancer

    DEFF Research Database (Denmark)

    Braidotti, P; Nuciforo, P G; Mollenhauer, J

    2004-01-01

    with anti DMBT1 antibody (DMBTh12) and sections from 36 samples, were double-stained also with anti MCM5, one of the 6 pre-replicative complex proteins with cell proliferation-licensing functions. DMBT1 gene expression at mRNA level was assessed by RT-PCR in frozen tissues samples from 39 patients. RESULTS...... expression was down-regulated in the cancerous lesions compared to the normal and/or hyperplastic epithelium adjacent to carcinomas (3/55 positive carcinomas versus 33/42 positive normal/hyperplastic epithelia; p = 0.0001). In 72% of cases RT-PCR confirmed immunohistochemical results. Most of normal...... and hyperplastic mammary cells positive with DMBTh12 were also MCM5-positive. CONCLUSIONS: The redistribution and up-regulation of DMBT1 in normal and hyperplastic tissues flanking malignant tumours and its down-regulation in carcinomas suggests a potential role in breast cancer. Moreover, the concomitant...

  16. miR-125b suppresses the proliferation and migration of osteosarcoma cells through down-regulation of STAT3

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Li-hong; Li, Hui; Li, Jin-ping; Zhong, Hui; Zhang, Han-chon; Chen, Jia [Department of Orthopedics, The Second Xiangya Hospital of Central South University, Changsha 410010 (China); Xiao, Tao, E-mail: xiaotaoxyl@163.com [Department of Orthopedics, The Second Xiangya Hospital of Central South University, Changsha 410010 (China)

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer miR-125b is frequently down-regulated in osteosarcoma samples and human osteosarcoma cell lines. Black-Right-Pointing-Pointer Ectopic restoration of miR-125b suppresses cell proliferation and migration in vitro. Black-Right-Pointing-Pointer STAT3 is the direct and functional downstream target of miR-125b. Black-Right-Pointing-Pointer STAT3 can bind to the promoter region of miR-125b and serves as a transactivator. -- Abstract: There is accumulating evidence that microRNAs are involved in multiple processes in development and tumor progression. Abnormally expressed miR-125b was found to play a fundamental role in several types of cancer; however, whether miR-125b participates in regulating the initiation and progress of osteosarcoma still remains unclear. Here we demonstrate that miR-125b is frequently down-regulated in osteosarcoma samples and human osteosarcoma cell lines. The ectopic restoration of miR-125b expression in human osteosarcoma cells suppresses proliferation and migration in vitro and inhibits tumor formation in vivo. We further identified signal transducer and activator of transcription 3 (STAT3) as the direct and functional downstream target of miR-125b. Interestingly, we discovered that the expression of miR-125b is regulated by STAT3 at the level of transcription. STAT3 binds to the promoter region of miR-125b in vitro and serves as a transactivator. Taken together, our findings point to an important role in the molecular etiology of osteosarcoma and suggest that miR-125b is a potential target in the treatment of osteosarcoma.

  17. Squamous Cell Carcinoma

    Science.gov (United States)

    ... Kids’ zone Video library Find a dermatologist Squamous cell carcinoma Overview Squamous cell carcinoma: This man's skin ... a squamous cell carcinoma on his face. Squamous cell carcinoma: Overview Squamous cell carcinoma (SCC) is a ...

  18. SOD2 deficient erythroid cells up-regulate transferrin receptor and down-regulate mitochondrial biogenesis and metabolism.

    Directory of Open Access Journals (Sweden)

    Florent M Martin

    Full Text Available BACKGROUND: Mice irradiated and reconstituted with hematopoietic cells lacking manganese superoxide dismutase (SOD2 show a persistent hemolytic anemia similar to human sideroblastic anemia (SA, including characteristic intra-mitochondrial iron deposition. SA is primarily an acquired, clonal marrow disorder occurring in individuals over 60 years of age with uncertain etiology. METHODOLOGY/PRINCIPAL FINDINGS: To define early events in the pathogenesis of this murine model of SA, we compared erythroid differentiation of Sod2⁻/⁻ and normal bone marrow cells using flow cytometry and gene expression profiling of erythroblasts. The predominant transcriptional differences observed include widespread down-regulation of mitochondrial metabolic pathways and mitochondrial biogenesis. Multiple nuclear encoded subunits of complexes I-IV of the electron transport chain, ATP synthase (complex V, TCA cycle and mitochondrial ribosomal proteins were coordinately down-regulated in Sod2⁻/⁻ erythroblasts. Despite iron accumulation within mitochondria, we found increased expression of transferrin receptor, Tfrc, at both the transcript and protein level in SOD2 deficient cells, suggesting deregulation of iron delivery. Interestingly, there was decreased expression of ABCb7, the gene responsible for X-linked hereditary SA with ataxia, a component required for iron-sulfur cluster biogenesis. CONCLUSIONS/SIGNIFICANCE: These results indicate that in erythroblasts, mitochondrial oxidative stress reduces expression of multiple nuclear genes encoding components of the respiratory chain, TCA cycle and mitochondrial protein synthesis. An additional target of particular relevance for SA is iron:sulfur cluster biosynthesis. By decreasing transcription of components of cluster synthesis machinery, both iron utilization and regulation of iron uptake are impacted, contributing to the sideroblastic phenotype.

  19. Srcasm down-regulates src family tyrosine kinases fyn expression in human esophageal squamohus cell carcinoma TE1 cell line%Srcasm负调控Src家族酪氨酸激酶Fyn在人食管鳞状细胞癌TE1细胞株中的表达

    Institute of Scientific and Technical Information of China (English)

    齐宇; 李鑫; 崔广晖; 胡伟; 赵松

    2010-01-01

    Objective To determine whether Fyn expression was altered in human esophageal squamous cell carcinoma (ESCC) TE1 cell line by TomL1. Methods We transfected 0, 0. 25, 0. 5,1.0, and 2.0 μg Srcasm plasmid DNA into TE1 cells and examined Fyn/Srcasm expression by Western blotting. Results Srcasm decreased the Fyn protein expression in TE1 cells in a dose-dependent manner.Conclusion Srcasm may act as a negative regulator of Fyn in human ESCC, and play an important role in human ESCC progress.%目的 观察人食管鳞状细胞癌TE1细胞株中Srcasm对Src家族酪氨酸激酶Fyn表达的影响.方法 分别转染0、0.25、0.5、1.0、2.0 μg人Srcasm质粒DNA入TE1细胞株中观察其对Fyn表达的影响.结果 转染Srcasm质粒DNA可以降低人食管鳞状细胞癌TE1细胞中Fyn的表达,且呈剂量依赖性.结论 人食管鳞状细胞癌中Srcasm可能是Src家族酪氨酸激酶Fyn的负调控因子,在食管鳞状细胞癌的发生发展过程中起重要作用.

  20. Topotecan inhibits cancer cell migration by down-regulation of chemokine CC motif receptor 7 and matrix metalloproteinases

    Institute of Scientific and Technical Information of China (English)

    Sen-sen LIN; Li SUN; Yan-kai ZHANG; Ren-ping ZHAO; Wen-lu LIANG; Sheng-tao YUAN; Lu-yong ZHANG

    2009-01-01

    Aim: The aim of this study was to investigate the effect of topotecan (TPT) on cancer cell migration.Methods: Growth inhibition of TPT was analyzed by MTT assay, and cancer cell migration was measured by transwell double chamber assay. To verify the effect of TPT on the chemokine receptors CXCR4 and CCR7, quantitative PCR, semi-quantitative PCR and Western blot analysis were performed. The secretion of MMP-2 and MMP-9 was detected by enzyme-linked immunosorbent assay (ELISA) and gelatin zymography. To evaluate possible contributions of CCR7 to MMP secretion, the overexpression vectors pcDNA3.1+-CCR7 and CCR7 siRNA were transiently transfected into MDA-MB-435 cells.Results: TPT inhibited cancer cell migration in a dose-dependent manner. Additionally, TPT significantly decreased the expression of CCR7 in both MDA-MB-435 and MDA-MB-231 cells and moderately reduced the expression of CXCR4 in MDA-MB-435 cells. The secretion of MMPs (MMP-2, MMP-9) was also inhibited by TPT. Overexpression of CCR7 increased the secretion of MMP-2/9 and cancer cell migration, whereas knockdown of CCR7 reduced active MMP-2/9 production and migration of MDA-MB-435 cells.Conclusion: TPT inhibited cancer cell migration by down-regulation of CCR7 and MMPs (MMP-2 and MMP-9).

  1. The Cell Surface Estrogen Receptor, G Protein- Coupled Receptor 30 (GPR30, is Markedly Down Regulated During Breast Tumorigenesis

    Directory of Open Access Journals (Sweden)

    Indira Poola

    2008-01-01

    Full Text Available Background: GPR30 is a cell surface estrogen receptor that has been shown to mediate a number of non-genomic rapid effects of estrogen and appear to balance the signaling of estrogen and growth factors. In addition, progestins appear to use GPR30 for their actions. Therefore, GPR30 could play a critical role in hormonal regulation of breast epithelial cell integrity. Deregulation of the events mediated by GPR30 could contribute to tumorigenesis.Methods: To understand the role of GPR30 in the deregulation of estrogen signaling processes during breast carcinogenesis, we have undertaken this study to investigate its expression at mRNA levels in tumor tissues and their matched normal tissues. We compared its expression at mRNA levels by RT quantitative real-time PCR relative to GAPDH in ERα”—positive (n = 54 and ERα”—negative (n = 45 breast cancer tissues to their matched normal tissues.Results: We report here, for the first time, that GPR30 mRNA levels were significantly down-regulated in cancer tissues in comparison with their matched normal tissues (p 0.0001 by two sided paired t-test. The GPR30 expression levels were significantly lower in tumor tissues from patients (n = 29 who had lymph node metastasis in comparison with tumors from patients (n = 53 who were negative for lymph node metastasis (two sample t-test, p 0.02, but no association was found with ERα, PR and other tumor characteristics.Conclusions: Down-regulation of GPR30 could contribute to breast tumorigenesis and lymph node metastasis.

  2. Down-Regulating Cold Shock Protein Genes Impairs Cancer Cell Survival and Enhances Chemosensitivity

    OpenAIRE

    2009-01-01

    The microenvironment of the cancer cell is pivotal to its phenotypic regulation. One of the central components of the microenvironment is temperature. An elevation in environmental temperature has been shown to increase the cancer cell's susceptibility to chemo- and radiation therapy. The goal of the studies described here was to identify some of the pathways that are modified by a mild increase in temperature in cancer cells. Using prostate cancer cells as a model system we found that in add...

  3. Down-regulation of LAK cell-mediated cytotoxicity: cancer and ageing.

    Science.gov (United States)

    Bykovskaya, S N; Abronina, I F; Kupriyanova, T A; Bubenik, J

    1990-01-01

    A comparative study was made of the generation of lymphokine-activated killer (LAK) cells in patients with melanoma and healthy donors of different age groups. Significant reduction of effector cell cytotoxicity in patients following 72 h culture with 1,000 U/ml or recombinant IL-2 (rIL-2) as well as a decreased ability to generate LAK cells in elderly individuals were shown to be correlated with suppressor cell activation in rIL-2 stimulated cell population. Suppressor effect depends on monocytes and T-lymphocytes: partial abolition of suppression in LAK cells was observed following removal of adherent cells or treatment with OKT8 monoclonal antibodies and complement.

  4. Down-regulation of β-catenin Nuclear Localization by Aspirin Correlates with Growth Inhibition of Jurkat Cell Line

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    In this study, we examined the effects of aspirin on the growth rates, subcellar distribution of β-catenin protein, the expression of β-catenin/TCF signaling pathway target gene cyclinD1 mRNA,and cell cycle of Jurkat cell line (Human T-acute lymphoblastic leukemia). Our results showed that the treatment with aspirin inhibited the growth of Jurkat cell line. Jurkat cells treated with 3 mmol/L of aspirin could significantly decrease nuclear localization of β-catenin, and at 5 mmol/L of aspirin,the nuclear localization of β-catenin was undetectable. QRT-PCR showed that the target gene cyclinD1 mRNA expression was gradually decreased with the dosage of aspirin. Aspirin induced G0/G1cell cycle arrest in Jurkat cells. We are led to conclude that aspirin acts through β-catenin-independent mechanisms. The effects of aspirin include down-regulation of β-catenin nuclear localization and G0/G1 cell cycle arrest, which might serve as a means of growth inhibition in aspirin-treated human Jurkat cell line.

  5. Berberine Induces Apoptosis in p53-Null Leukemia Cells by Down-Regulating XIAP at the Post-Transcriptional Level

    Directory of Open Access Journals (Sweden)

    Jian Liu

    2013-11-01

    Full Text Available Background: Berberine exerts anticancer activities both in vitro and in vivo through different mechanisms. However, the underlying molecular mechanisms of berberine induced p53-independent apoptosis remain unclear. Methods: The p53-null leukemia cell line EU-4 cells were exposed to berberine. Then the cell viability and apoptosis were determined. Western blot and PCR were employed to detect the expression of apoptosis related protein, XIAP and MDM2. Small interfering RNA (siRNA was applied to knock down endogenous expression of MDM2 and XIAP. Results: Berberine induced p53-independent, XIAP-mediated apoptotic cell death in p53-null leukemia cells. Treatment with berberine resulted in suppression of XIAP protein in a dose- and time- dependent manner. Berberine induced down-regulation of XIAP protein involving inhibition of MDM2 expression and a proteasome-dependent pathway. Moreover, inhibition of XIAP by berberine or siRNA increased the sensitivity of leukemia cells to doxorubicin-induced apoptosis. Conclusion: Our findings characterize the molecular mechanisms of berberine-induced caspase activation and subsequent apoptosis, and berberine may be a novel candidate inducer of apoptosis in leukemia cells, which normally lack p53 expression.

  6. Cisplatin and ultra-violet-C synergistically down-regulate receptor tyrosine kinases in human colorectal cancer cells

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    Kawaguchi Junji

    2012-07-01

    Full Text Available Abstract Background Platinum-containing anti-cancer drugs such as cisplatin are widely used for patients with various types of cancers, however, resistance to cisplatin is observed in some cases. Whereas we have recently reported that high dose UV-C (200 J/m² induces colorectal cancer cell proliferation by desensitization of EGFR, which leads oncogenic signaling in these cells, in this study we investigated the combination effect of low dose cisplatin (10 μM and low dose UV-C (10 J/m² on cell growth and apoptosis in several human colorectal cancer cells, SW480, DLD-1, HT29 and HCT116. Results The combination inhibited cell cycle and colony formation, while either cisplatin or UV-C alone had little effect. The combination also induced apoptosis in these cells. In addition, the combination caused the downregulation of EGFR and HER2. Moreover, UV-C alone caused the transient internalization of the EGFR, but with time EGFR recycled back to the cell surface, while cisplatin did not affect its localization. Surprisingly, the combination caused persistent internalization of the EGFR, which results in the lasting downregulation of the EGFR. Conclusions The combination of low dose cisplatin and low dose UV-C synergistically exerted anti-cancer effect by down-regulating RTK, such as EGFR and HER2. These findings may provide a novel strategy for the treatment of patients with colorectal cancer.

  7. Cimetidine down-regulates stability of Foxp3 protein via Stub1 in Treg cells.

    Science.gov (United States)

    Zhang, Yizhi; Chen, Zhoujia; Luo, Xuerui; Wu, Bin; Li, Bin; Wang, Bin

    2016-10-02

    Foxp3-expressing Treg cells have been well documented to provide immune regulation by promoting immune tolerance and suppressing immune over-reaction. Cimetidine (CIM), used to inhibit stomach acid secretion, has been reported to promote immune responses and suppress Treg cell function in several studies. However, the underlying mechanism is unknown. To investigate CIM effects on the suppressive function of Treg and Foxp3, here we used CIM to stimulate human CD4(+)CD25(+) Treg cells and Jurkat T cells and evaluated changes of Foxp3 expression and stability. Our data showed that CIM leads to a reduction of Foxp3 via E3 ligase Stub1-mediated proteosomal degradation, which is dependent on an activated PI3K-AKT-mTOR pathway. Thus, CIM affects the suppressive function of Treg cells by destabilizing their Foxp3 expression.

  8. Induction of leukemia cell apoptosis by cheliensisin A involves down-regulation of Bcl-2 expression

    Institute of Scientific and Technical Information of China (English)

    Li ZHONG; Chao-ming LI; Xiao-jiang HAO; Li-guang LOU

    2005-01-01

    Aim: To investigate the apoptosis-inducing effect of cheliensisin A (GC-51), a novel styryl-lactone isolated from Goniothalamus cheliensis, on human promyelocytic leukemia HL-60 cells and the mechanism of action involved.Methods: Apoptotic cell death was determined by morphological examination and DNA agarose gel electrophoresis. The activity of caspase-3 was assessed using Western blotting and the expression of Bcl-2 and Bax genes was analyzed using the reverse transcription-polymerase chain reaction (RT-PCR) method. Results:GC-51 significantly inhibited the proliferation of HL-60 cells with an IC50 of 2.4±0.2 μmol/L and effectively induced apoptosis in HL-60 cells. Exposure of HL-60cells to 10 μmol/L GC-51 for 8 h resulted in approximately 53% of the cells under going apoptosis. Caspase-3 was activated in GC-51-treated cells, which was manifested by the appearance of the 17 kDa active form of caspase-3 and the cleavage of poly(ADP-ribose) polymerase (PARP). Meanwhile, GC-51 markedly reduced the expression of the anti-apoptotic gene Bcl-2 and increased the expression of the pro-apoptotic gene Bax. The apoptosis-inducing effect of GC-51 was cAMP dependent protein kinase (PKA) dependent because PKA, but not the protein kinase C, specific inhibitor H-89, blocked the induction of apoptosis by GC-51 in HL-60 cells. Conclusion: The results demonstrate that GC-51 effectively induces apoptosis in HL-60 cells and that this effect is PKA-dependent and involves the downregulation of Bcl-2 expression and the activation of caspase-3.

  9. MicroRNA-224 inhibits proliferation and migration of breast cancer cells by down-regulating Fizzled 5 expression.

    Science.gov (United States)

    Liu, Feng; Liu, Yang; Shen, Jingling; Zhang, Guoqiang; Han, Jiguang

    2016-08-02

    The Wnt/β-catenin signaling is crucial for the proliferation and migration of breast cancer cells. However, the expression of microRNA-224 (miR-224) in the different types of breast cancers and its role in the Wnt/β-catenin signaling and the proliferation and migration of breast cancer cells are poorly understood. In this study, the levels of miR-224 in different types of breast cancer tissues and cell lines were examined by quantitative RT-PCR and the potential targets of miR-224 in the Wnt/β-catenin signaling were investigated. The effects of altered miR-224 expression on the frequency of CD44+CD24- cancer stem-like cells (CSC), proliferation and migration of MCF-7 and MDA-MB-231 cells were examined by flow cytometry, MTT and transwell migration. We found that the levels of miR-224 expression in different types of breast cancer tissues and cell lines were associated inversely with aggressiveness of breast cancers. Enhanced miR-224 expression significantly reduced the fizzled 5-regulated luciferase activity in 293T cells, fizzled 5 expression in MCF-7 and MDA-MB-231 cells, the β-dependent luciferase activity in MCF-7 cells, and the nuclear translocation of β-catenin in MDA-MB-231 cells. miR-224 inhibition significantly increased the percentages of CSC in MCF-7 cells and enhanced proliferation and migration of MCF-7 cells. Enhanced miR-224 expression inhibited proliferation and migration of MDA-MB-231 cells, and the growth of implanted breast cancers in vivo. Induction of Frizzled 5 over-expression mitigated the miR-224-mediated inhibition of breast cancer cell proliferation. Collectively, these data indicated that miR-224 down-regulated the Wnt/β-catenin signaling possibly by binding to Frizzled 5 and inhibited proliferation and migration of breast cancer cells.

  10. Overexpressed ubiquitin ligase Cullin7 in breast cancer promotes cell proliferation and invasion via down-regulating p53

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Hongsheng [Department of Histology and Embryology, Guangdong Medical College, Dongguan 523808, Guangdong (China); Wu, Fenping [The 7th People’s Hospital of Chengdu, Chengdu 610041, Sichuan (China); Wang, Yan [The Second School of Clinical Medicine, Guangdong Medical College, Dongguan 523808, Guangdong (China); Yan, Chong [School of Pharmacy, Guangdong Medical College, Dongguan 523808, Guangdong (China); Su, Wenmei, E-mail: wenmeisutg@126.com [Oncology of Affiliated Hospital Guangdong Medical College, Zhanjiang 524000, Guangdong (China)

    2014-08-08

    Highlights: • Cullin7 is overexpressed in human breast cancer samples. • Cullin7 stimulated proliferation and invasion of breast cancer cells. • Inhibition of p53 contributes to Cullin7-induced proliferation and invasion. - Abstract: Ubiquitin ligase Cullin7 has been identified as an oncogene in some malignant diseases such as choriocarcinoma and neuroblastoma. However, the role of Cullin7 in breast cancer carcinogenesis remains unclear. In this study, we compared Cullin7 protein levels in breast cancer tissues with normal breast tissues and identified significantly higher expression of Cullin7 protein in breast cancer specimens. By overexpressing Cullin7 in breast cancer cells HCC1937, we found that Cullin7 could promote cell growth and invasion in vitro. In contrast, the cell growth and invasion was inhibited by silencing Cullin7 in breast cancer cell BT474. Moreover, we demonstrated that Cullin7 promoted breast cancer cell proliferation and invasion via down-regulating p53 expression. Thus, our study provided evidence that Cullin7 functions as a novel oncogene in breast cancer and may be a potential therapeutic target for breast cancer management.

  11. Galectin-1 is expressed in early-type neural progenitor cells and down-regulates neurogenesis in the adult hippocampus

    Directory of Open Access Journals (Sweden)

    Imaizumi Yoichi

    2011-01-01

    Full Text Available Abstract Background In the adult mammalian brain, neural stem cells (NSCs proliferate in the dentate gyrus (DG of the hippocampus and generate new neurons throughout life. A multimodal protein, Galectin-1, is expressed in neural progenitor cells (NPCs and implicated in the proliferation of the NPCs in the DG. However, little is known about its detailed expression profile in the NPCs and functions in adult neurogenesis in the DG. Results Our immunohistochemical and morphological analysis showed that Galectin-1 was expressed in the type 1 and 2a cells, which are putative NSCs, in the subgranular zone (SGZ of the adult mouse DG. To study Galectin-1's function in adult hippocampal neurogenesis, we made galectin-1 knock-out mice on the C57BL6 background and characterized the effects on neurogenesis. In the SGZ of the galectin-1 knock-out mice, increased numbers of type 1 cells, DCX-positive immature progenitors, and NeuN-positive newborn neurons were observed. Using triple-labeling immunohistochemistry and morphological analyses, we found that the proliferation of the type-1 cells was increased in the SGZ of the galectin-1 knock-out mice, and we propose that this proliferation is the mechanism for the net increase in the adult neurogenesis in these knock-out mice DG. Conclusions Galectin-1 is expressed in the neural stem cells and down-regulates neurogenesis in the adult hippocampus.

  12. KCNK5 is Functionally Down-Regulated Upon Long-Term Hypotonicity in Ehrlich Ascites Tumor Cells

    DEFF Research Database (Denmark)

    Kirkegaard, S. S.; Wulff, Tune; Gammeltoft, S.;

    2013-01-01

    Background/Aims: Regulatory volume decrease (RVD) in response to acute cell swelling is well described and KCNK5 (also known as TASK-2 or K2P5.1) has been shown to be the volume sensitive K+ channel in Ehrlich cells. Very little is, on the other hand, known about the effects of long-term hypotoni......Background/Aims: Regulatory volume decrease (RVD) in response to acute cell swelling is well described and KCNK5 (also known as TASK-2 or K2P5.1) has been shown to be the volume sensitive K+ channel in Ehrlich cells. Very little is, on the other hand, known about the effects of long......-term hypotonicity on expression and function of KCNK5, thus we have investigated the effect of long-term hypotonicity (24h - 48h) on KCNK5 in Ehrlich cells on the mRNA, protein and physiological levels. Methods: Physiological effects of long-term hypotonicity were measured using patch-clamp and Coulter counter...... physiological impairment of KCNK5 in Ehrlich cells after long-term hypotonic stimulation is predominantly due to down-regulation of the KCNK5 protein synthesis.© 2013 S. Karger AG, Basel...

  13. Impact of transient down-regulation of DREAM in human embryonic stem cell pluripotency

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    A. Fontán-Lozano

    2016-05-01

    Full Text Available Little is known about the functions of downstream regulatory element antagonist modulator (DREAM in embryonic stem cells (ESCs. However, DREAM interacts with cAMP response element-binding protein (CREB in a Ca2+-dependent manner, preventing CREB binding protein (CBP recruitment. Furthermore, CREB and CBP are involved in maintaining ESC self-renewal and pluripotency. However, a previous knockout study revealed the protective function of DREAM depletion in brain aging degeneration and that aging is accompanied by a progressive decline in stem cells (SCs function. Interestingly, we found that DREAM is expressed in different cell types, including human ESCs (hESCs, human adipose-derived stromal cells (hASCs, human bone marrow-derived stromal cells (hBMSCs, and human newborn foreskin fibroblasts (hFFs, and that transitory inhibition of DREAM in hESCs reduces their pluripotency, increasing differentiation. We stipulate that these changes are partly mediated by increased CREB transcriptional activity. Overall, our data indicates that DREAM acts in the regulation of hESC pluripotency and could be a target to promote or prevent differentiation in embryonic cells.

  14. Moderate Hypoxia Down-Regulates Interleukin-6 Secretion and TLR4 Expression in Human Sw.71 Placental Cells

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    Koumei Shirasuna

    2015-07-01

    Full Text Available Background/Aims: The placenta is a vital organ for pregnancy. Many in vitro placental experiments are conducted under 21% O2; however, O2 tension could influence cellular functions, including cytokine secretion. We investigated the effects of oxygen tension between moderate hypoxia (5% O2 and normoxia (21% O2 by testing the hypothesis that moderate hypoxia regulates cellular phenotypes differently from normoxia in human trophoblast cells. Methods and Results: Sw.71 trophoblast cells were incubated under normoxic or moderately hypoxic conditions. Cells were also treated with lipopolysaccharide (LPS as a Toll-like receptor 4 (TLR4 ligand inducing inflammation. Interleukin-6 (IL-6 as an inflammatory cytokine was determined, and TLR4, hypoxia-induced factor-1α (HIF1α, and reactive oxygen species (ROS production were detected. Moderate hypoxia increased HIF1α expression and cell proliferation and acted by two different mechanisms to decrease IL-6 secretion compared with normoxia: it limits the TLR4 expression and ROS production. Treatment with cobalt chloride as an HIF1 activator inhibited IL-6 secretion and TLR4 expression; this effect was reversed on treatment with PX-12 as an HIF1 suppressor. Conclusion: IL-6 secretion, TLR4 expression, and ROS production, classical markers of inflammation, are down-regulated by moderate hypoxia, and HIF1α and ROS have a potential to regulate these responses in human trophoblast cells.

  15. KCNK5 is Functionally Down-Regulated Upon Long-Term Hypotonicity in Ehrlich Ascites Tumor Cells

    Directory of Open Access Journals (Sweden)

    Signe Skyum Kirkegaard

    2013-11-01

    Full Text Available Background/Aims: Regulatory volume decrease (RVD in response to acute cell swelling is well described and KCNK5 (also known as TASK-2 or K2P5.1 has been shown to be the volume sensitive K+ channel in Ehrlich cells. Very little is, on the other hand, known about the effects of long-term hypotonicity on expression and function of KCNK5, thus we have investigated the effect of long-term hypotonicity (24h - 48h on KCNK5 in Ehrlich cells on the mRNA, protein and physiological levels. Methods: Physiological effects of long-term hypotonicity were measured using patch-clamp and Coulter counter techniques. Expression patterns of KCNK5 on mRNA and protein levels were established using real-time qPCR and western blotting respectively. Results: The maximum swelling-activated current through KCNK5 was significantly decreased upon 48h of hypotonicity and likewise the RVD response was significantly impaired after both 24 and 48h of hypotonic stimulation. No significant differences in the KCNK5 mRNA expression patterns between control and stimulated cells were observed, but a significant decrease in the KCNK5 protein level 48h after stimulation was found. Conclusion: The data suggest that the strong physiological impairment of KCNK5 in Ehrlich cells after long-term hypotonic stimulation is predominantly due to down-regulation of the KCNK5 protein synthesis.

  16. Effects of Notch-1 down-regulation on malignant behaviors of breast cancer stem cells.

    Science.gov (United States)

    Peng, Gong-ling; Tian, Ye; Lu, Chong; Guo, Hui; Zhao, Xiang-wang; Guo, Ya-wen; Wang, Long-qiang; Du, Qiu-li; Liu, Chun-ping

    2014-04-01

    This study examined the effect of Notch-1 signaling on malignant behaviors of breast cancer cells by regulating breast cancer stem cells (BCSCs). BCSCs were enriched by using serum-free medium and knocked out of Notch-1 by using a lentiviral vector. Real-time polymerase chain reaction (RT-PCR) and Western blotting were used to detect the Notch-1 expression levels in breast cancer cell lines and BCSCs, and flow cytometry to detect the proportion of BCSCs in BCSC spheres. The BCSC self-renewal, migration, invasion, and tumorigenicity were examined by the tumor microsphere-forming assay and transwell assay and after xenotransplantation. The results showed that the Notch-1 silencing reduced the number of BCSC spheres, the proportion of BCSCs, and the number of cells penetrating through the transwell membrane. It also decreased the size of tumors that were implanted in the nude mice. These results suggest that Notch-1 signaling is intimately linked to the behaviors of BCSCs. Blocking Notch-1 signaling can inhibit the malignant behaviors of BCSCs, which may provide a promising therapeutical approach for breast cancer.

  17. Effects of Down-regulation of Integrin-β_1 Expression on Migration and Hepatic Metastasis of Human Colon Carcinoma

    Institute of Scientific and Technical Information of China (English)

    张建立; 高军; 谭晓杰; 王敏; 秦仁义

    2010-01-01

    Organ-specific tumor cell adhesion to extracellular matrix (ECM) components and cell migration into host organs often involve integrin-mediated cellular processes. Direct integrin-mediated cell adhesion to ECM components in the space of Disse appears to be required for the successful liver metastatic formation of colon cancer. In the present study, human colon cancer HT-29 cells were transfected by liposome with integrin-β1 antisense oligodeoxynucleotide (ASODN). The integrin-β1 gene expression in HT-29 cel...

  18. BCL-2 family protein, BAD is down-regulated in breast cancer and inhibits cell invasion

    Energy Technology Data Exchange (ETDEWEB)

    Cekanova, Maria, E-mail: mcekanov@utk.edu [Department of Small Animal Clinical Sciences, College of Veterinary Medicine, The University of Tennessee, Knoxville, TN (United States); Fernando, Romaine I. [Department of Obstetrics and Gynecology, Graduate School of Medicine, Medical Center, The University of Tennessee, Knoxville, TN (United States); Siriwardhana, Nalin [Department of Animal Science, The University of Tennessee, Knoxville, TN (United States); Sukhthankar, Mugdha [Department of Biomedical and Diagnostics Sciences, College of Veterinary Medicine, The University of Tennessee, Knoxville, TN (United States); Parra, Columba de la [Department of Biochemistry, School of Medicine, University of Puerto Rico, Medical Sciences Campus, San Juan, PR (United States); Woraratphoka, Jirayus [Department of Obstetrics and Gynecology, Graduate School of Medicine, Medical Center, The University of Tennessee, Knoxville, TN (United States); Malone, Christine [Laboratory of Molecular Carcinogenesis, National Institute of Environmental Health Sciences, Research Triangle Park, NC (United States); Ström, Anders [Center for Nuclear Receptors and Cell Signaling, Department of Biology and Biochemistry, University of Houston, Houston, TX (United States); Baek, Seung J. [Department of Biomedical and Diagnostics Sciences, College of Veterinary Medicine, The University of Tennessee, Knoxville, TN (United States); Wade, Paul A. [Laboratory of Molecular Carcinogenesis, National Institute of Environmental Health Sciences, Research Triangle Park, NC (United States); Saxton, Arnold M. [Department of Animal Science, The University of Tennessee, Knoxville, TN (United States); Donnell, Robert M. [Department of Biomedical and Diagnostics Sciences, College of Veterinary Medicine, The University of Tennessee, Knoxville, TN (United States); Pestell, Richard G. [Department of Cancer Biology, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA (United States); and others

    2015-02-01

    We have previously demonstrated that the anti-apoptotic protein BAD is expressed in normal human breast tissue and shown that BAD inhibits expression of cyclin D1 to delay cell-cycle progression in breast cancer cells. Herein, expression of proteins in breast tissues was studied by immunohistochemistry and results were analyzed statistically to obtain semi-quantitative data. Biochemical and functional changes in BAD-overexpressing MCF7 breast cancer cells were evaluated using PCR, reporter assays, western blotting, ELISA and extracellular matrix invasion assays. Compared to normal tissues, Grade II breast cancers expressed low total/phosphorylated forms of BAD in both cytoplasmic and nuclear compartments. BAD overexpression decreased the expression of β-catenin, Sp1, and phosphorylation of STATs. BAD inhibited Ras/MEK/ERK and JNK signaling pathways, without affecting the p38 signaling pathway. Expression of the metastasis-related proteins, MMP10, VEGF, SNAIL, CXCR4, E-cadherin and TlMP2 was regulated by BAD with concomitant inhibition of extracellular matrix invasion. Inhibition of BAD by siRNA increased invasion and Akt/p-Akt levels. Clinical data and the results herein suggest that in addition to the effect on apoptosis, BAD conveys anti-metastatic effects and is a valuable prognostic marker in breast cancer. - Highlights: • BAD and p-BAD expressions are decreased in breast cancer compared with normal breast tissue. • BAD impedes breast cancer invasion and migration. • BAD inhibits the EMT and transcription factors that promote cancer cell migration. • Invasion and migration functions of BAD are distinct from the BAD's role in apoptosis.

  19. BCL-2 family protein, BAD is down-regulated in breast cancer and inhibits cell invasion.

    Science.gov (United States)

    Cekanova, Maria; Fernando, Romaine I; Siriwardhana, Nalin; Sukhthankar, Mugdha; De la Parra, Columba; Woraratphoka, Jirayus; Malone, Christine; Ström, Anders; Baek, Seung J; Wade, Paul A; Saxton, Arnold M; Donnell, Robert M; Pestell, Richard G; Dharmawardhane, Suranganie; Wimalasena, Jay

    2015-02-01

    We have previously demonstrated that the anti-apoptotic protein BAD is expressed in normal human breast tissue and shown that BAD inhibits expression of cyclin D1 to delay cell-cycle progression in breast cancer cells. Herein, expression of proteins in breast tissues was studied by immunohistochemistry and results were analyzed statistically to obtain semi-quantitative data. Biochemical and functional changes in BAD-overexpressing MCF7 breast cancer cells were evaluated using PCR, reporter assays, western blotting, ELISA and extracellular matrix invasion assays. Compared to normal tissues, Grade II breast cancers expressed low total/phosphorylated forms of BAD in both cytoplasmic and nuclear compartments. BAD overexpression decreased the expression of β-catenin, Sp1, and phosphorylation of STATs. BAD inhibited Ras/MEK/ERK and JNK signaling pathways, without affecting the p38 signaling pathway. Expression of the metastasis-related proteins, MMP10, VEGF, SNAIL, CXCR4, E-cadherin and TlMP2 was regulated by BAD with concomitant inhibition of extracellular matrix invasion. Inhibition of BAD by siRNA increased invasion and Akt/p-Akt levels. Clinical data and the results herein suggest that in addition to the effect on apoptosis, BAD conveys anti-metastatic effects and is a valuable prognostic marker in breast cancer.

  20. Allele loss and down-regulation of heparanase gene are associated with the progression and poor prognosis of hepatocellular carcinoma.

    Directory of Open Access Journals (Sweden)

    Guo-Liang Huang

    Full Text Available OBJECTIVES: The role of heparanase (HPSE gene in cancers including hepatocellular carcinoma (HCC is currently controversial. This study was aimed at investigating the impact of genetic alteration and expression change of HPSE on the progression and prognosis of HCC. METHODS: The HPSE gene was studied in three different aspects: (1 loss of heterozygosity (LOH by a custom SNP microarray and DNA copy number by real-time PCR; (2 mRNA level by qRT-PCR; and (3 protein expression by immunohistochemistry. The clinical significances of allele loss and expression change of HPSE were analyzed. RESULTS: Microarray analysis showed that the average LOH frequency for 10 SNPs located within HPSE gene was 31.6%, three of which were significantly correlated with tumor grade, serum HBV-DNA level, and AFP concentration. In agreement with SNP LOH data, DNA copy number loss of HPSE was observed in 38.74% (43/111 of HCC cases. HPSE mRNA level was notably reduced in 74.1% (83/112 of tumor tissues compared with non-tumor liver tissues, which was significantly associated with DNA copy number loss, increased tumor size, and post-operative metastasis. HPSE protein level was also remarkably reduced in 66.3% (53/80 of tumor tissues, which was correlated with tumor grade. Patients with lower expression level of HPSE mRNA or protein had a significantly lower survival rate than those with higher expression. Cox regression analysis suggested that HPSE protein was an independent predictor of overall survival in HCC patients. CONCLUSIONS: The results in this study demonstrate that genetic alteration and reduction of HPSE expression are associated with tumor progression and poor prognosis of HCCs, suggesting that HPSE behaves like a tumor suppressor gene and is a potential prognostic marker for HCC patients.

  1. Down-regulation of NF-κB signaling by Gordonia bronchialis prevents the activation of gut epithelial cells.

    Science.gov (United States)

    Smaldini, Paola L; Stanford, John; Romanin, David E; Fossati, Carlos A; Docena, Guillermo H

    2014-08-01

    The immunomodulatory power of heat-killed Gordonia bronchialis was studied on gut epithelial cells activated with pro-inflammatory stimuli (flagellin, TNF-α or IL-1β). Light emission of luciferase-transfected epithelial cells and mRNA expression of IL-1β, TNF-α, IL-6, CCL20, IL-8 and MCP-1 were measured. NF-κB activation was assessed by immunofluorescence and immunoblotting, and induction of reactive oxygen species (ROS) was evaluated. In vivo inhibitory properties of G. bronchialis were studied with ligated intestinal loop assay and in a mouse model of food allergy. G. bronchialis promoted the down-regulation of the expression of CCL20 and IL-1β on activated epithelial cells in a dose-dependent manner. A concomitant blocking of nuclear p65 translocation with increased production of ROS was found. In vivo experiments confirmed the inhibition of CCL20 expression and the suppression of IgE sensitization and hypersensitivity symptoms in the food allergy mouse model. In conclusion, heat-killed G. bronchialis inhibited the activation of NF-κB pathway in human epithelial cells, and suppressed the expression of CCL20. These results indicate that G. bronchialis may be used to modulate the initial steps of innate immune activation, which further suppress the allergic sensitization. This approach may be exploited as a therapy for intestinal inflammation.

  2. VEGFR3 Inhibition Chemosensitizes Ovarian Cancer Stemlike Cells through Down-Regulation of BRCA1 and BRCA2

    Directory of Open Access Journals (Sweden)

    Jaeyoung Lim

    2014-04-01

    Full Text Available In ovarian cancer, loss of BRCA gene expression in tumors is associated with improved response to chemotherapy and increased survival. A means to pharmacologically downregulate BRCA gene expression could improve the outcomes of patients with BRCA wild-type tumors. We report that vascular endothelial growth factor receptor 3 (VEGFR3 inhibition in ovarian cancer cells is associated with decreased levels of both BRCA1 and BRCA2. Inhibition of VEGFR3 in ovarian tumor cells was associated with growth arrest. CD133+ ovarian cancer stemlike cells were preferentially susceptible to VEGFR3-mediated growth inhibition. VEGFR3 inhibition–mediated down-regulation of BRCA gene expression reversed chemotherapy resistance and restored chemosensitivity in resistant cell lines in which a BRCA2 mutation had reverted to wild type. Finally, we demonstrate that tumor-associated macrophages are a primary source of VEGF-C in the tumor microenvironment. Our studies suggest that VEGFR3 inhibition may be a pharmacologic means to downregulate BRCA genes and improve the outcomes of patients with BRCA wild-type tumors.

  3. Resveratrol down-regulates survivin and induces apoptosis in human multidrug-resistant SPC-A-1/CDDP cells.

    Science.gov (United States)

    Zhao, Weiguo; Bao, Pengtao; Qi, Haowen; You, Houcheng

    2010-01-01

    We studied the effect of resveratrol treatment on multidrug-resistant human non-small cell lung cancer cells. Human multidrug-resistant SPC-A-1/CDDP cells were treated with resveratrol at a concentration of 25, 50, or 100 microM in in vitro studies and nude mice were implanted with multidrug-resistant SPC-A-1/and fed a special diet that included resveratrol at a dose of either 1 g/kg/day or 3 g/kg/day in in vivo studies. No adverse toxicological effects of resveratrol treatment were observed. The rate of cell proliferation, apoptosis ratio, cell cycle phase distribution, IC50 values of cisplatin, gefitinib, and paclitaxel, implanted tumour volume, and expression of survivin in resveratrol-treated and control mice were then determined. Resveratrol significantly inhibited the proliferation of SPC-A-1/CDDP cells, induced apoptosis, arrested the cell cycle phase between G0-G1 and S phase or at the G2/M phase, decreased the IC50 values of multiple chemotherapeutic drugs, and showed anti-tumour effects in nude mice that had been implanted with SPC-A-1/CDDP cells. In additional, resveratrol affected the proliferation of SPC-A-1/CDDP cells in a dose- and time-dependent manner. Expression of survivin in SPC-A-1/CDDP cells decreased after they were treated with all concentrations of resveratrol and resveratrol was also found to have a dose-dependent effect on survivin expression. Resveratrol can induce apoptosis in multidrug-resistant human NSCLC SPC-A-1/CDDP cells by down-regulating the expression of survivin.

  4. Infant intestinal Enterococcus faecalis down-regulates inflammatory responses in human intestinal cell lines

    Institute of Scientific and Technical Information of China (English)

    Shugui Wang; Lydia Hui Mei Ng; Wai Ling Chow; Yuan Kun Lee

    2008-01-01

    AIM:To investigate the ability of Lactic acid bacteria (LAB)to modulate inflammatory reaction in human intestinal celllines(Caco-2,HT-29 and HCT 116).Different strains of LAB isolatedfrom new born infants and fermented milk,together withthestrains obtained from culture collectionsweretested.METHODS:LABs were treated with human intestinal cell lines.ELISA was used to detect IL-8 and TGF-β protein secretion.Cytokines and Toll like receptors (TLRs) gene expression were assessed using RT-PCR.Conditional medium,sonicated bacteria and UV killed bacteria were used to find the effecter molecules on the bacteria.Carbohydrate oxidation and protein digestion were applied to figure out the molecules'residues.Adhesion assays were further carried out.RESULTS:It was found that Enterococcus faecalis is the main immune modulator among the LABs by downregulation of IL-8 secretion and upregulation of TGF-β.Strikingly,the effect was only observed in four strains of E.faecalis out of the 27 isolated and tested.This implies strain dependent immunomodulation in the host.In addition,E.faecalis may regulate inflammatory responses through TLR3,TLR4,TLR9 and TRAF6.Carbohydrates on the bacterial cell surface are involved in both its adhesion to intestinal cells and regulation of inflammatory responses in the host.CONCLUSION:These data provide a case for the modulation of intestinal mucosal immunity in which specific strains of E.faecalis have uniquely evolved to maintain colonic homeostasis and regulate inflammatoryresponses.

  5. miR-449 inhibits cell proliferation and is down-regulated in gastric cancer

    Directory of Open Access Journals (Sweden)

    Federspiel Birgitte

    2011-03-01

    Full Text Available Abstract Background Gastric cancer is the fourth most common cancer in the world and the second most prevalent cause of cancer related death. The development of gastric cancer is mainly associated with H. Pylori infection leading to a focus in pathology studies on bacterial and environmental factors, and to a lesser extent on the mechanistic development of the tumour. MicroRNAs are small non-coding RNA molecules involved in post-transcriptional gene regulation. They are found to regulate genes involved in diverse biological functions and alterations in microRNA expression have been linked to the pathogenesis of many malignancies. The current study is focused on identifying microRNAs involved in gastric carcinogenesis and to explore their mechanistic relevance by characterizing their targets. Results Invitrogen NCode miRNA microarrays identified miR-449 to be decreased in 1-year-old Gastrin KO mice and in H. Pylori infected gastric tissues compared to tissues from wild type animals. Growth rate of gastric cell lines over-expressing miR-449 was inhibited by 60% compared to controls. FACS cell cycle analysis of miR-449 over-expressing cells showed a significant increase in the sub-G1 fraction indicative of apoptosis. ß-Gal assays indicated a senescent phenotype of gastric cell lines over-expressing miR-449. Affymetrix 133v2 arrays identified GMNN, MET, CCNE2, SIRT1 and CDK6 as miR-449 targets. Luciferase assays were used to confirm GMNN, MET, CCNE2 and SIRT1 as direct targets. We also show that miR-449 over-expression activated p53 and its downstream target p21 as well as the apoptosis markers cleaved CASP3 and PARP. Importantly, qPCR analyses showed a loss of miR-449 expression in human clinical gastric tumours compared to normal tissues. Conclusions In this study, we document a diminished expression of miR-449 in Gastrin KO mice and further confirmed its loss in human gastric tumours. We investigated the function of miR-449 by identifying its

  6. miR-214 down-regulates ARL2 and suppresses growth and invasion of cervical cancer cells.

    Science.gov (United States)

    Peng, Ruiqing; Men, Jianlong; Ma, Rui; Wang, Qian; Wang, Yang; Sun, Ying; Ren, Jing

    2017-03-11

    Increasing evidence has shown that miRNAs are implicated in carcinogenesis and can function as oncogenes or tumor suppressor genes in human cancers. In this study, we confirmed that miR-214 is frequently down-regulated in cervical cancer compared with normal cervical tissues. Ectopic expression of miR-214 suppressed proliferation, migration and invasion of HeLa and C33A cervical cancer cells. Bioinformatics analysis revealed that ADP ribosylation factor like 2 (ARL2) was a potential target of miR-214 and was remarkably up-regulated in cervical cancer. Knockdown of ARL2 markedly inhibited cervical cancer cell proliferation, migration and invasion, similarly to over-expression of miR-214, indicating that ARL2 may function as an oncogene in cervical cancer. In conclusion, our study revealed that miR-214 acts as a tumor suppressor via inhibiting proliferation, migration and invasion of cervical cancer cells through targeting ARL2, and that both miR-214 and ARL2 may serve as prognostic or therapeutic targets for cervical cancer.

  7. Bone marrow mesenchymal stromal cells affect the cell cycle arrest effect of genotoxic agents on acute lymphocytic leukemia cells via p21 down-regulation.

    Science.gov (United States)

    Zhang, Yiran; Hu, Kaimin; Hu, Yongxian; Liu, Lizhen; Wang, Binsheng; Huang, He

    2014-09-01

    The effect of bone marrow microenvironment on the cell cycle of acute lymphocytic leukemia (ALL) and the underlying mechanism has not been elucidated. In this study, we found that in normal condition, bone marrow mesenchymal stromal cells (BM-MSCs) had no significant effect on the cell cycle and apoptosis of ALL; in the condition when the cell cycle of ALL was blocked by genotoxic agents, BM-MSCs could increase the S-phase cell ratio and decrease the G2/M phase ratio of ALL. Besides, BM-MSCs could protect ALL cells from drug-induced apoptosis. Then, we proved that BM-MSCs affect the cell cycle arrest effect of genotoxic agents on ALL cells via p21 down-regulation. Moreover, our results indicated that activation of Wnt/β-catenin and Erk pathways might be involved in the BM-MSC-induced down-regulation of p21 in ALL cells. Targeting microenvironment-related signaling pathway may therefore be a potential novel approach for ALL therapy.

  8. The excreted polysaccharide of Pleurotus eryngii inhibits the foam-cell formation via down-regulation of CD36.

    Science.gov (United States)

    Chen, Jingjing; Yong, Yangyang; Xia, Xian; Wang, Zeliang; Liang, Youxing; Zhang, Shizhu; Lu, Ling

    2014-11-04

    Previous study has verified the polysaccharide from the fruiting body of Pleurotus eryngii (PEPE) is capable of decreasing the lipid content in both of cell-line and mouse model. However, little is known about underlying mechanisms and whether this bioactive polysaccharide exists in submerged culture. Here, we verified the excreted polysaccharides EP and EP-1 from submersion culture of P. eryngii have the remarkable inhibitory effects on lipid accumulation in macrophage-derived foam cells. Structure analysis indicates EP-1 consists of D-types of glucose, galactose and mannose with the main β(1 → 3)-glucan glycosidic linkage branched at O-6 by α-D-glucose while EP digested by β-1,3-glucanase fails to decrease the lipid accumulation, suggesting that the special structure is essential for its function. Expression analysis suggests that EP is able to cause the down-regulation of the scavenger receptor-CD36 on both transcription and protein levels. Most importantly, EP can be obtained by fermentation in a mass-production.

  9. eEF-2 Phosphorylation Down-Regulates P-Glycoprotein Over-Expression in Rat Brain Microvessel Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Xing Hua Tang

    Full Text Available We investigated whether glutamate, NMDA receptors, and eukaryote elongation factor-2 kinase (eEF-2K/eEF-2 regulate P-glycoprotein expression, and the effects of the eEF-2K inhibitor NH125 on the expression of P-glycoprotein in rat brain microvessel endothelial cells (RBMECs.Cortex was obtained from newborn Wistar rat brains. After surface vessels and meninges were removed, the pellet containing microvessels was resuspended and incubated at 37°C in culture medium. Cell viability was assessed by the MTT assay. RBMECs were identified by immunohistochemistry with anti-vWF. P-glycoprotein, phospho-eEF-2, and eEF-2 expression were determined by western blot analysis. Mdr1a gene expression was analyzed by RT-PCR.Mdr1a mRNA, P-glycoprotein and phospho-eEF-2 expression increased in L-glutamate stimulated RBMECs. P-glycoprotein and phospho-eEF-2 expression were down-regulated after NH125 treatment in L-glutamate stimulated RBMECs.eEF-2K/eEF-2 should have played an important role in the regulation of P-glycoprotein expression in RBMECs. eEF-2K inhibitor NH125 could serve as an efficacious anti-multidrug resistant agent.

  10. Down-regulation of heat shock protein HSP90ab1 in radiation-damaged lung cells other than mast cells.

    Science.gov (United States)

    Haase, Michael G; Geyer, Peter; Fitze, Guido; Baretton, Gustavo B

    2014-05-01

    Ionizing radiation (IR) leads to fibrosing alveolitis (FA) after a lag period of several weeks to months. In a rat model, FA starts at 8 weeks after IR. Before that, at 5.5 weeks after IR, the transcription factors Sp1 (stimulating protein 1) and AP-1 (activator protein 1) are inactivated. To find genes/proteins that were down-regulated at that time, differentially expressed genes were identified in a subtractive cDNA library and verified by quantitative RT-PCR (reverse transcriptase polymerase chain reaction), western blotting and immunohistochemistry (IH). The mRNA of the molecular chaperone HSP90AB1 (heat shock protein 90 kDa alpha, class B member 1) was down-regulated 5.5 weeks after IR. Later, when FA manifested, HSP90ab1 protein was down-regulated by more than 90% in lung cells with the exception of mast cells. In most mast cells of the normal lung, both HSP90ab1 and HSP70, another major HSP, show a very low level of expression. HSP70 was massively up-regulated in all mast cells three months after irradiation whereas HSP90AB1 was up-regulated only in a portion of mast cells. The strong changes in the expression of central molecular chaperones may contribute to the well-known disturbance of cellular functions in radiation-damaged lung tissue.

  11. Apigenin suppresses migration and invasion of transformed cells through down-regulation of C-X-C chemokine receptor 4 expression

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Lei; Kuang, Lisha; Hitron, John Andrew; Son, Young-Ok; Wang, Xin; Budhraja, Amit [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States); Lee, Jeong-Chae [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States); Institute of Oral Biosciences and BK21 Program, Research Center of Bioactive Materials, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Pratheeshkumar, Poyil [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States); Chen, Gang [Department of Internal Medicine, University of Kentucky College of Medicine, Lexington, KY 40536 (United States); Zhang, Zhuo [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States); Luo, Jia [Department of Internal Medicine, University of Kentucky College of Medicine, Lexington, KY 40536 (United States); Shi, Xianglin, E-mail: xshi5@email.uky.edu [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States)

    2013-10-01

    Environmental exposure to arsenic is known to cause various cancers. There are some potential relationships between cell malignant transformation and C-X-C chemokine receptor type 4 (CXCR4) expressions. Metastasis, one of the major characteristics of malignantly transformed cells, contributes to the high mortality of cells. CXCR4 and its natural chemokine ligand C-X-C motif ligand 12 (CXCL12) play a critical role in metastasis. Therefore, identification of nutritional factors which are able to inhibit CXCR4 is important for protection from environmental arsenic-induced carcinogenesis and for abolishing metastasis of malignantly transformed cells. The present study demonstrates that apigenin (4′,5,7-trihydroxyflavone), a natural dietary flavonoid, suppressed CXCR4 expression in arsenic-transformed Beas-2B cells (B-AsT) and several other types of transformed/cancer cells in a dose- and time-dependent manner. Neither proteasome nor lysosome inhibitor had any effect in reducing the apigenin-induced down-regulation of CXCR4, indicating that apigenin-induced down-regulation of CXCR4 is not due to proteolytic degradation. The down-regulation of CXCR4 is mainly due to the inhibition of nuclear factor κB (NF-κB) transcriptional activity. Apigenin also abolished migration and invasion of transformed cells induced by CXCL12. In a xenograft mouse model, apigenin down-regulated CXCR4 expression and suppressed tumor growth. Taken together, our results show that apigenin is a novel inhibitor of CXCR4 expression. This dietary flavonoid has the potential to suppress migration and invasion of transformed cells and prevent environmental arsenic-induced carcinogenesis. - Highlights: • Apigenin has a potential in preventing environmental arsenic induced carcinogenesis. • Apigenin suppresses CXCR4 in malignant transformed cells in vitro and in vivo. • The down-regulation of CXCR4 is mainly due to inhibition of NF-κB activity.

  12. Transcriptional down regulation of hTERT and senescence induction in HepG2 cells by chelidonine

    Institute of Scientific and Technical Information of China (English)

    Sakineh Kazemi Noureini; Michael Wink

    2009-01-01

    aged with large volume and dark staining in the β-galactosidase assay. CONCLUSION: Chelidonine reduces telomerase activity through down-regulation of hTERT expression. Senescence induction might not be directly caused by reducing telomerase activity as it occurs after a few population doublings.

  13. Down-regulation of miR-133a as a poor prognosticator in non-small cell lung cancer.

    Science.gov (United States)

    Wang, Yuzhou; Li, Jinmei; Chen, Hongming; Mo, Yanli; Ye, Haiyin; Luo, Yiping; Guo, Kangwen; Mai, Zongjiong; Zhang, Ying; Chen, Baoying; Zhou, Yijin; Yang, Zhixiong

    2016-10-15

    miR-133a has been demonstrated to play an important role in tumor progression. The aim of present study was to analyze the correlation between miR-133a expression level and clinicopathologic features and its prognostic significance in non-small cell lung cancer (NSCLC). The expression of miR-133a in 104 pairs of human lung cancer tissues and adjacent normal lung tissues were analyzed by qRT-PCR. Here we show that miR-133a was down-regulated in NSCLC. The levels of miR-133a were negatively correlated with the status of N classification (N0-N1 vs. N2-N3, P=0.000), clinical stage (I-II vs. III-IV, P=0.010) and MMP-14 expression (High vs. Low, P=0.012). The patients with low miR-133a expression had shorter survival time than those with high miR-133a expression. Multivariate analysis indicated that the level of miR-133a expression was an independent prognostic indicator (P=0.012) for the survival of patients with NSCLC. In conclusion, decreased expression of miR-133a might be a potential unfavorable prognostic factor for patients with NSCLC, and further studies would be needed to prove our findings.

  14. Basal Cell Carcinoma

    Science.gov (United States)

    ... Kids’ zone Video library Find a dermatologist Basal cell carcinoma Overview Basal cell carcinoma: This skin cancer ... that has received years of sun exposure. Basal cell carcinoma: Overview Basal cell carcinoma (BCC) is the ...

  15. Atherosclerosis-Associated Endothelial Cell Apoptosis by MiR-429-Mediated Down Regulation of Bcl-2

    Directory of Open Access Journals (Sweden)

    Tao Zhang

    2015-10-01

    -associated endothelial cell apoptosis may result from down regulation of Bcl-2, through increased miR-429 that binds and suppresses translation of Bcl-2 mRNA.

  16. Internalization and down-regulation of mu opioid receptors by endomorphins and morphine in SH-SY5Y human neuroblastoma cells.

    Science.gov (United States)

    Horner, Kristen A; Zadina, James E

    2004-12-03

    The human neuroblastoma cell line, SH-SY5Y, was used to examine the effects of morphine and the endogenous opioid peptides, endomorphin-1 (EM-1) and endomorphin-2 (EM-2), on mu opioid receptor (MOR) internalization and down-regulation. Treatment for 24 h with EM-1, EM-2 or morphine at 100 nM, 1 microM and 10 microM resulted in a dose-dependent down-regulation of mu receptors. Exposure of cells to 10 microM EM-1 for 2.5, 5 and 24 h resulted in a time-dependent down-regulation of mu receptors. Down-regulation of mu receptors by morphine and EM-1 was blocked by treatment with hypertonic sucrose, consistent with an endocytosis-dependent mechanism. Sensitive cell-surface binding studies with a radiolabeled mu antagonist revealed that morphine was able to induce internalization of mu receptors naturally expressed in SH-SY5Y cells. EM-1 produced a more rapid internalization of mu receptors than morphine, but hypertonic sucrose blocked the internalization induced by each of these agonists. This study demonstrates that, like morphine, the endomorphins down-regulate mu opioid receptors in a dose- and time-dependent manner. This study also demonstrates that morphine, as well as EM-1, can induce rapid, endocytosis-dependent internalization of mu opioid receptors in SH-SY5Y cells. These results may help elucidate the ability of mu agonists to regulate the number and responsiveness of their receptors.

  17. Metformin suppresses CYP1A1 and CYP1B1 expression in breast cancer cells by down-regulating aryl hydrocarbon receptor expression

    Energy Technology Data Exchange (ETDEWEB)

    Do, Minh Truong; Kim, Hyung Gyun; Tran, Thi Thu Phuong; Khanal, Tilak; Choi, Jae Ho [Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon (Korea, Republic of); Chung, Young Chul [Department of Food Science and Culinary, International University of Korea, Jinju (Korea, Republic of); Jeong, Tae Cheon, E-mail: taecheon@ynu.ac.kr [College of Pharmacy, Yeungnam University, Gyeongsan (Korea, Republic of); Jeong, Hye Gwang, E-mail: hgjeong@cnu.ac.kr [Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon (Korea, Republic of)

    2014-10-01

    Induction of cytochrome P450 (CYP) 1A1 and CYP1B1 by environmental xenobiotic chemicals or endogenous ligands through the activation of the aryl hydrocarbon receptor (AhR) has been implicated in a variety of cellular processes related to cancer, such as transformation and tumorigenesis. Here, we investigated the effects of the anti-diabetes drug metformin on expression of CYP1A1 and CYP1B1 in breast cancer cells under constitutive and inducible conditions. Our results indicated that metformin down-regulated the expression of CYP1A1 and CYP1B1 in breast cancer cells under constitutive and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced conditions. Down-regulation of AhR expression was required for metformin-mediated decreases in CYP1A1 and CYP1B1 expression, and the metformin-mediated CYP1A1 and CYP1B1 reduction is irrelevant to estrogen receptor α (ERα) signaling. Furthermore, we found that metformin markedly down-regulated Sp1 protein levels in breast cancer cells. The use of genetic and pharmacological tools revealed that metformin-mediated down-regulation of AhR expression was mediated through the reduction of Sp1 protein. Metformin inhibited endogenous AhR ligand-induced CYP1A1 and CYP1B1 expression by suppressing tryptophan-2,3-dioxygenase (TDO) expression in MCF-7 cells. Finally, metformin inhibits TDO expression through a down-regulation of Sp1 and glucocorticoid receptor (GR) protein levels. Our findings demonstrate that metformin reduces CYP1A1 and CYP1B1 expression in breast cancer cells by down-regulating AhR signaling. Metformin would be able to act as a potential chemopreventive agent against CYP1A1 and CYP1B1-mediated carcinogenesis and development of cancer. - Graphical abstract: Schematic of the CYP1A1 and CYP1B1 gene regulation by metformin. - Highlights: • Metformin inhibits CYP1A1 and CYP1B1 expression. • Metformin down-regulates the AhR signaling. • Metformin reduces Sp1 protein expression. • Metformin suppresses TDO expression.

  18. siRNA against presenilin 1 (PS1 down regulates amyloid β42 production in IMR-32 cells

    Directory of Open Access Journals (Sweden)

    Kandimalla Ramesh JL

    2012-01-01

    Full Text Available Abstract Background One of the pathological hallmarks of Alzheimer's disease (AD is the deposition of the ~4 kDa amyloid β protein (Aβ within lesions known as senile plaques. Aβ is also deposited in the walls of cerebral blood vessels in many cases of AD. A substantial proportion of the Aβ that accumulates in the AD brain is deposited as Amyloid, which is highly insoluble, proteinaceous material with a β-pleated-sheet conformation and deposited extracellularly in the form of 5-10 nm wide straight fibrils. As γ-secretase catalyzes the final cleavage that releases the Aβ42 or 40 from amyloid β -protein precursor (APP, therefore, it is a potential therapeutic target for the treatment of AD. γ-Secretase cleavage is performed by a high molecular weight protein complex containing presenilins (PSs, nicastrin, Aph-1 and Pen-2. Previous studies have demonstrated that the presenilins (PS1 and PS2 are critical components of a large enzyme complex that performs γ-secretase cleavage. Methods In this study we used RNA interference (RNAi technology to examine the effects of small-interfering RNA (siRNA against PS1 on expression levels of PS1 and Aβ42 in IMR-32 Cells using RTPCR, western blotting and immunofluorescence techniques. Results The results of the present study showed down regulation of PS1 and Aβ42 in IMR32 cells transfected with siRNA against PS1. Conclusion Our results substantiate the concept that PS1 is involved in γ-secretase activity and provides the rationale for therapeutic strategies aimed at influencing Aβ42 production.

  19. Hemoglobin A1c induced down-regulation of CD36 of Plasmodium Falciparum parasitized red cell

    Directory of Open Access Journals (Sweden)

    Hassan Hijazi, Atif Alagib, Hisham Waggiallah

    2014-03-01

    Full Text Available Objective: High values of glycosylated hemoglobin have been found to correlate with decreased deformability of erythrocyte. CD36 (Cluster of Differentiation 36 is an integral membrane protein found on the surface of many cell types of class B scavenger receptor family. Plasmodium falciparum and diabetes mellitus is associated many complications. Aim of this study to investigate the down-regulation of HbA1c to CD36 on P. falciparum parasitized red blood cells Diabetes mellitus patients. Methods: This is cross section study conducted among diabetic patients attending in Jabir Abo Eleiz diabetic center in Khartoum state. Venous blood samples were collected in heparin containers for Plasmodium falciparum culture, and random blood sugar. For HbA1c in 0.04 mg EDTA anticoagulant, 2-5 ml of blood was collected. Sample size was 45 samples and was collected from known diabetic patients with HbA1c more than 8%. All data were analyzed by using Statistical Package for Social Science (SPSS. Results: show the mean difference between CD36 negative control and CD36 positive control was found to be statistically significant increasing of CD36 presence at P. value =0.001 (P ≤0.001. The mean difference between CD36 positive control and diabetic patients with HbA1C more 8% was found to be statistically significant reduction of CD36 expression at p=0.001. Conclusion: Hyperglycemia (HbA1c leads to decrease of CD36 expression and interfere with innate and active immunity. In this study HbA1c participates in increasing of P. falciparum malaria complications.

  20. MiR-128b is down-regulated in gastric cancer and negatively regulates tumour cell viability by targeting PDK1/Akt/NF-κB axis

    Indian Academy of Sciences (India)

    Ling Zhang; Jun Lei; Zi-Ling Fang; Jian-Ping Xiong

    2016-03-01

    Gastric cancer (GC) is the fourth most prevalent type of cancer worldwide, which is usually caused by the interaction between environmental and genetic factors, or epigenetic aspects. Referring to the non-coding RNAs, miR-128b has been reported to be associated with many tumour cases, and exerts distinct functions in different types of cancers. However, the function of miR-128b in GC onset and progression largely remains unknown. In the present study, we found that miR-128b expression was down-regulated in tissues from 18 GC patients and 3 carcinoma cell lines. In turn, over-expression of miR-128b suppressed GC cell proliferation, invasion and promoted apoptosis. Moreover, miR-128b was predicted to bind the 3'UTR of PDK1 gene using bioinformatic target-screening tools. Accordingly, ectopic expression of miR-128b inhibited the PDK1 expression at both transcriptional and post-transcriptional levels, and furthermore, the expression of gene tailed by the 3′UTR of PDK1 gene was significantly decreased in a dualluciferase reporter assay, suggesting that PDK1 was a direct target of miR-128b in GC cells. In the conditon of miR-128b over-expression, we also observed spontaneous inactivation of the Akt/NF-κB signalling, implying PDK1 was a potential regulator of this pathway. In conclusion, our study shed some novel light on miR-128b-PDK1/Akt/NF-κB axis on GC progression.

  1. Down-regulated peroxisome proliferator-activated receptor γ (PPARγ) in lung epithelial cells promotes a PPARγ agonist-reversible proinflammatory phenotype in chronic obstructive pulmonary disease (COPD).

    Science.gov (United States)

    Lakshmi, Sowmya P; Reddy, Aravind T; Zhang, Yingze; Sciurba, Frank C; Mallampalli, Rama K; Duncan, Steven R; Reddy, Raju C

    2014-03-07

    Chronic obstructive pulmonary disease (COPD) is a progressive inflammatory condition and a leading cause of death, with no available cure. We assessed the actions in pulmonary epithelial cells of peroxisome proliferator-activated receptor γ (PPARγ), a nuclear hormone receptor with anti-inflammatory effects, whose role in COPD is largely unknown. We found that PPARγ was down-regulated in lung tissue and epithelial cells of COPD patients, via both reduced expression and phosphorylation-mediated inhibition, whereas pro-inflammatory nuclear factor-κB (NF-κB) activity was increased. Cigarette smoking is the main risk factor for COPD, and exposing airway epithelial cells to cigarette smoke extract (CSE) likewise down-regulated PPARγ and activated NF-κB. CSE also down-regulated and post-translationally inhibited the glucocorticoid receptor (GR-α) and histone deacetylase 2 (HDAC2), a corepressor important for glucocorticoid action and whose down-regulation is thought to cause glucocorticoid insensitivity in COPD. Treating epithelial cells with synthetic (rosiglitazone) or endogenous (10-nitro-oleic acid) PPARγ agonists strongly up-regulated PPARγ expression and activity, suppressed CSE-induced production and secretion of inflammatory cytokines, and reversed its activation of NF-κB by inhibiting the IκB kinase pathway and by promoting direct inhibitory binding of PPARγ to NF-κB. In contrast, PPARγ knockdown via siRNA augmented CSE-induced chemokine release and decreases in HDAC activity, suggesting a potential anti-inflammatory role of endogenous PPARγ. The results imply that down-regulation of pulmonary epithelial PPARγ by cigarette smoke promotes inflammatory pathways and diminishes glucocorticoid responsiveness, thereby contributing to COPD pathogenesis, and further suggest that PPARγ agonists may be useful for COPD treatment.

  2. Down regulation of pRb in cultures of avian neuroretina cells promotes proliferation of reactive Müller-like cells and emergence of retinal stem/progenitors.

    Science.gov (United States)

    Marx, Maria; Lebuhotel, Céline; Laugier, Danielle; Chapelle, Audrey; Calothy, Georges; Saule, Simon

    2010-06-01

    The aim of this work was to define the role of pRb depletion in the proliferation and differentiation of avian retinoblasts in vitro. For this purpose vectors expressing pRb short hairpin RNA were used to deplete pRb in cultures of avian neuroretinal cells. Down regulation of pRb was observed by Western blot and quantification of nuclear pRb. Cell proliferation and differentiation were studied following BrdU labeling and immunostaining. Transfection significantly down-regulated pRb in neuroretinal cells. Long-term effect of pRb depletion mainly induced proliferation of epithelial-like cells that expressed markers of reactive Müller glial cells. A minority of these cells that survived passaging could be maintained as neurosphere-like aggregates with low pRb, not observed in control cultures. BrdU labeling followed by a two week chase showed the presence of cells still remained labelled, indicating low cell cycling. Under appropriate conditions, these aggregates differentiate in precursors of amacrine interneurons shown by the expression of AP2, in absence of the photoreceptors marker visinin and the late neuronal marker MAP2. Taken together these data show that decrease pRb level in cultures of avian neuroretinal cells promotes the emergence and proliferation of stem cell/progenitors from reactive-like Muller cells.

  3. Sulindac sulfide induces autophagic death in gastric epithelial cells via survivin down-regulation: a mechanism of NSAIDs-induced gastric injury.

    Science.gov (United States)

    Chiou, Shiun-Kwei; Hoa, Neil; Hodges, Amy

    2011-06-01

    Sulindac sulfide, a nonsteroidal anti-inflammatory drug (NSAID), has anti-tumorigenic and anti-inflammatory activities, but causes gastric mucosal damage. NSAIDs cause gastric injury in part by down-regulation of Survivin, an apoptosis inhibitor, resulting in apoptosis induction. Autophagy is a process that promotes cellular health by destroying unwanted cellular materials. Excessive autophagy induction could lead to a non-apoptotic cell death (autophagic cell death). The present study showed that sulindac sulfide at a physiological concentration also induces autophagic death in human gastric epithelial AGS and rat gastric epithelial RGM-1 cells, and that Survivin down-regulation is a mechanism involved: Sulindac sulfide treatment increased LC3b-II and APG7 levels and cytosolic vacuole formation, indications of autophagy induction, in AGS and RGM-1 cells. Sulindac sulfide treatment induced AGS and RGM-1 cell death, which was significantly reduced by pretreatment with the autophagy inhibitors 3-methyladenine and chloroquine, indicating that sulindac sulfide induced autophagic cell death. Stable overexpression of Survivin in RGM-1 cells did not inhibit the induction of LC3b-II levels or vacuole formation by sulindac sulfide, but significantly reduced the resulting cell death, suggesting that Survivin may inhibit autophagic cell death downstream of LC3b-II induction and vacuole formation. Indeed, siRNA depletion of LC3b in AGS cells inhibited the down-regulation of Survivin levels and the induction of cell death by sulindac sulfide, confirming that down-regulation of Survivin occurs in the autophagy pathway downstream of LC3b-II induction by sulindac sulfide. Induction of Survivin-dependent autophagic cell death is a novel mechanism by which sulindac sulfide induces gastric mucosal injury.

  4. 下调肝星状细胞 MACC1对胃癌细胞迁移侵袭能力的影响%Effect of down -regulating MACC1 by shRNA on invasion and migration of gastric carci-noma cells

    Institute of Scientific and Technical Information of China (English)

    袁园; 朱正秋

    2017-01-01

    Objective:By in vitro experiment to explore the regulatory effect of MACC1 gene silencing in HSC expressα-SMA,MMP -2,MMP -9 and HGF,and its effects on ability of migration and invasion of co -cultured gastric carcinoma cells.Methods:To construct targeting MACC1 gene eukaryotic expression interference plasmid (MACC1 -shRNA)and non -specific irrelevant interference plasmid (shNC),transient transfect them into HSC with lipofectamine -2000,named low -expression group(shRNA)and negative control group (NC)accordingly,non-transfected HSC treated as a blank control group(Blank -Ctrl).The above three groups of cells were cultured in accordance with routinel,the expression levels of MACC1,α-SMA,HGF,MMP -2 and MMP -9 in human hepatic stellate cells were detected by Western blot and RT -PCR.Three groups of cells were co -cultured with MGC803 gastric carcinoma cells in vitro,the capabilities of migration and invasion were measured by transwell assay.Results:Western -Blot results showed that the expression of MACC1,α-SMA,HGF,MMP -2 and MMP -9 protein in HSC was significantly lower than that in the negative group and blank control group after transfection of MACC1 -shRNA (P 0.05).Conclusion:Silencing MACC1 in hepatic stellate cells, can inhibit the expression of MMP -2,MMP -9 and HGF,and decrease the migration and invasion of gastric cancer cells in co -culture.%目的:通过体外实验探讨肝星状细胞(hepatic stellate cell,HSC)MACC1基因沉默后调节肝星状细胞激活标志物α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)、肝细胞生长因子(hepatocyte growth fac-tor,HGF)、基质金属蛋白酶2(matrix metalloproteinase -2,MMP -2)、基质金属蛋白酶9(matrix metalloprotein-ase -9,MMP -9)的表达,以及对共培养胃癌细胞迁移侵袭能力的影响。方法:利用 RNA 干扰技术,构建针对MACC1基因的真核表达干涉质粒(MACC1-shRNA)、非特异性无关

  5. Src family kinase inhibitor PP2 efficiently inhibits cervical cancer cell proliferation through down-regulating phospho-Src-Y416 and phospho-EGFR-Y1173.

    Science.gov (United States)

    Kong, Lu; Deng, Zhihong; Shen, Haiying; Zhang, Yuxiang

    2011-02-01

    Tyrosine (Y) kinases inhibitors have been approved for targeted treatment of cancer. However, their clinical use is limited to some cancers and the mechanism of their action remains unclear. Previous study has indicated that PP2, a selective inhibitor of the Src family of non-receptor tyrosine kinases (nRTK), efficiently repressed cervical cancer growth in vitro and in vivo. In this regard, our aims are to explore the mechanism of PP2 on cervical cancer cell growth inhibition by investigating the suppressive divergence among PP1, PP2, and a negative control compound PP3. MTT results showed that three compounds had different inhibitory effects on proliferation of two cervical cancer cells, HeLa and SiHa, and PP2 was most efficient in a time- and dose-dependent manner. Moreover, we found 10 μM PP2 down-regulated pSrc-Y416 (P < 0.05), pEGFR-Y845 (P < 0.05), and -Y1173 (P < 0.05) expression levels, while 10 μM PP1 down-regulated pSrc-Y416 (P < 0.05) and pEGFR-Y845 (P < 0.05), but not pEGFR-Y1173; 10 μM PP3 down-regulated only pEGFR-Y1173 (P < 0.05). PP2 could modulate cell cycle arrest by up-regulating p21(Cip1) and p27(Kip1) in both HeLa and SiHa cells and down-regulating expression of cyclin A, and cyclin dependent kinase-2, -4 (Cdk-2, -4) in HeLa and of cyclin B and Cdk-2 in SiHa. Our results indicate that Src pathway and EGFR pathway play different roles in the proliferation of cervical cancer cells and PP2 efficiently reduces cervical cancer cell proliferation by reduction of both Src and EGFR activity.

  6. Warfarin and coumarin-like Murraya paniculata extract down-regulate EpCAM-mediated cell adhesion: individual components versus mixture for studying botanical metastatic chemopreventives.

    Science.gov (United States)

    Shao, Jingwei; Zhou, Suxia; Jiang, Zhou; Chi, Ting; Ma, Ji; Kuo, Minliang; Lee, Alan Yueh-Luen; Jia, Lee

    2016-08-02

    We recently defined cancer metastatic chemoprevention as utilizing safe and effective molecules to comprehensively prevent the spark of activation-adhesion-extravasation-proliferation metastatic cascade caused by circulating tumor cells (CTCs). The strategy focuses on preventing the most important starting point of the cascade. We identified an extract from a well-known medical plant Murraya paniculata, which inhibited both embryonic implantation to human endometrium as traditionally-used for abortion and CTC adhesion to human endothelium. Here, we separated and characterized five coumarin-containing components (Z1-Z5) from the botanic extract. Flow cytometry revealed that within 1-100 μg/mL, Z3 and Z5 down-regulated EpCAM expression in human colon HCT116, whereas, Z1 and Z2 did oppositely. Warfarin and Z1-Z5 component mixture (CM) also down-regulated EpCAM expression. The down-regulation of EpCAM by Z3, Z5, CM and warfarin was confirmed by western blotting, and caused inhibition on adhesion of cancer cells to human endothelial cells. Rat coagulation study showed that warfarin prolonged prothrombin time, whereas, Z3 did not. The present studies revealed that, for the first time, warfarin and coumarin-like components Z3, Z5 and CM from Murraya paniculata could directly inhibit EpCAM-mediated cell-cell adhesion.

  7. HCV core protein-induced down-regulation of microRNA-152 promoted aberrant proliferation by regulating Wnt1 in HepG2 cells.

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    Shifeng Huang

    Full Text Available BACKGROUND: Hepatitis C virus (HCV has been reported to regulate cellular microRNAs (miRNAs. The HCV core protein is considered to be a potential oncoprotein in HCV-related hepatocellular carcinoma (HCV-HCC, but HCV core-regulated miRNAs are largely unknown. Our preliminary experiments revealed significant down-regulation of microRNA-152 (miR-152 by HCV core protein in HepG2 cells. Through target gene prediction softwares, Wnt1 was predicted to be a potential target of miR-152. The present study was initiated to investigate whether miR-152 is aberrantly regulated by the HCV core protein, and involved in the regulation of the aberrant proliferation of HCV-HCC cells. METHODS: MiR-152 levels were examined by stem-loop real-time RT-PCR (SLqRT-PCR. Cell proliferation was analyzed by MTT and colony formation assay. Cell cycle analysis was performed by flow cytometry. Luciferase reporter assay was conducted to confirm miRNA-target association. Wnt1 expression was determined by real-time qPCR and Western blotting. RESULTS: HCV core protein significantly suppressed miR-152 expression, and led to significant Wnt1 up-regulation with a concomitant aberrantly promoted proliferation. Moreover, we validated that miR-152 inhibition promoted, while miR-152 mimics inhibited cell proliferation. Using, qRT-PCR and western blot, Wnt1 was demonstrated to be regulated by miR-152. Luciferase activity assay showed that while miR-152 mimics significantly reduced the luciferase activity by 83.76% (P<0.0001, miR-152 inhibitor showed no effect on luciferase reporter. Most notably, salvage expression of miR-152 after Ad-HCV core infection for 24 h almost totally reversed the proliferation-promoting effect of the HCV core protein, and meanwhile, reduced the expression of both Wnt1 mRNA and protein to basal levels. CONCLUSION: These findings provide important evidence that the reduced miR-152 expression by HCV core protein can indirectly lose an inhibitory effect on Wnt1

  8. AS1411-Induced Growth Inhibition of Glioma Cells by Up-Regulation of p53 and Down-Regulation of Bcl-2 and Akt1 via Nucleolin

    OpenAIRE

    Cheng, Ye; Zhao, Gang; Zhang, Siwen; Nigim, Fares; Zhou, Guangtong; Yu, Zhiyun; Song, Yang; Chen, Yong; Li, Yunqian

    2016-01-01

    AS1411 binds nucleolin (NCL) and is the first oligodeoxynucleotide aptamer to reach phase I and II clinical trials for the treatment of several cancers. However, the mechanisms by which AS1411 targets and kills glioma cells and tissues remain unclear. Here we report that AS1411 induces cell apoptosis and cycle arrest, and inhibits cell viability by up-regulation of p53 and down-regulation of Bcl-2 and Akt1 in human glioma cells. NCL was overexpressed in both nucleus and cytoplasm in human gli...

  9. Expression of the IL-7 receptor alpha-chain is down regulated on the surface of CD4 T-cells by the HIV-1 Tat protein.

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    Denny McLaughlin

    Full Text Available HIV infection elicits defects in CD4 T-cell homeostasis in both a quantitative and qualitative manner. Interleukin-7 (IL-7 is essential to T-cell homeostasis and several groups have shown reduced levels of the IL-7 receptor alpha-chain (CD127 on both CD4 and CD8 T-cells in viremic HIV+ patients. We have shown previously that soluble HIV Tat protein specifically down regulates cell surface expression of CD127 on human CD8 T-cells in a paracrine fashion. The effects of Tat on CD127 expression in CD4 T-cells has yet to be described. To explore this effect, CD4 T-cells were isolated from healthy individuals and expression levels of CD127 were examined on cells incubated in media alone or treated with Tat protein. We show here that, similar to CD8 T-cells, the HIV-1 Tat protein specifically down regulates CD127 on primary human CD4 T-cells and directs the receptor to the proteasome for degradation. Down regulation of CD127 in response to Tat was seen on both memory and naive CD4 T-cell subsets and was blocked using either heparin or anti-Tat antibodies. Tat did not induce apoptosis in cultured primary CD4 T-cells over 72 hours as determined by Annexin V and PI staining. Pre-incubation of CD4 T-cells with HIV-1 Tat protein did however reduce the ability of IL-7 to up regulate Bcl-2 expression. Similar to exogenous Tat, endogenously expressed HIV Tat protein also suppressed CD127 expression on primary CD4 T-cells. In view of the important role IL-7 plays in lymphocyte proliferation, homeostasis and survival, down regulation of CD127 by Tat likely plays a central role in immune dysregulation and CD4 T-cell decline. Understanding this effect could lead to new approaches to mitigate the CD4 T-cell loss evident in HIV infection.

  10. SKP2 siRNA inhibits the degradation of P27kip1 and down-regulates the expression of MRP in HL-60/A cells.

    Science.gov (United States)

    Xiao, Jie; Yin, Songmei; Li, Yiqing; Xie, Shuangfeng; Nie, Danian; Ma, Liping; Wang, Xiuju; Wu, Yudan; Feng, Jianhong

    2009-08-01

    S-phase kinase-associated protein 2 (SKP2) gene is a tumor suppressor gene, and is involved in the ubiquitin-mediated degradation of P27kip1. SKP2 and P27kip1 affect the proceeding and prognosis of leukemia through regulating the proliferation, apoptosis and differentiation of leukemia cells. In this study, we explored the mechanism of reversing of HL-60/A drug resistance through SKP2 down-regulation. HL-60/A cells were nucleofected by Amaxa Nucleofector System with SKP2 siRNA. The gene and protein expression levels of Skp2, P27kip1, and multi-drug resistance associated protein (MRP) were determined by reverse transcription-polymerase chain reaction and western blot analysis, respectively. The cell cycle was analyzed by flow cytometry. The 50% inhibitory concentration value was calculated using cytotoxic analysis according to the death rate of these two kinds of cells under different concentrations of chemotherapeutics to compare the sensitivity of the cells. HL-60/A cells showed multi-drug resistance phenotype characteristic by cross-resistance to adriamycin, daunorubicin, and arabinosylcytosine, due to the expression of MRP. We found that the expression of SKP2 was higher in HL-60/A cells than in HL-60 cells, but the expression of P27kip1 was lower. The expression of SKP2 in HL-60/A cells nucleofected by SKP2 siRNA was down-regulated whereas the protein level of P27kip1 was up-regulated. Compared with the MRP expression level in the control group (nucleofected by control siRNA), the mRNA and protein expression levels of MRP in HL-60/A cells nucleofected by SKP2 siRNA were lower, and the latter cells were more sensitive to adriamycin, daunorubicin, and arabinosylcytosine. Down-regulating the SKP2 expression and arresting cells in the G0/G1 phase improve drug sensitivity of leukemia cells with down-regulated MRP expression.

  11. AS1411-Induced Growth Inhibition of Glioma Cells by Up-Regulation of p53 and Down-Regulation of Bcl-2 and Akt1 via Nucleolin.

    Science.gov (United States)

    Cheng, Ye; Zhao, Gang; Zhang, Siwen; Nigim, Fares; Zhou, Guangtong; Yu, Zhiyun; Song, Yang; Chen, Yong; Li, Yunqian

    2016-01-01

    AS1411 binds nucleolin (NCL) and is the first oligodeoxynucleotide aptamer to reach phase I and II clinical trials for the treatment of several cancers. However, the mechanisms by which AS1411 targets and kills glioma cells and tissues remain unclear. Here we report that AS1411 induces cell apoptosis and cycle arrest, and inhibits cell viability by up-regulation of p53 and down-regulation of Bcl-2 and Akt1 in human glioma cells. NCL was overexpressed in both nucleus and cytoplasm in human glioma U87, U251 and SHG44 cells compared to normal human astrocytes (NHA). AS1411 bound NCL and inhibited the proliferation of glioma cells but not NHA, which was accompanied with up-regulation of p53 and down-regulation of Bcl-2 and Akt1. Moreover, AS1411 treatment resulted in the G2/M cell cycle arrest in glioma cells, which was however abolished by overexpression of NCL. Further, AS1411 induced cell apoptosis, which was prevented by silencing of p53 and overexpression of Bcl-2. In addition, AS1411 inhibited the migration and invasion of glioma cells in an Akt1-dependent manner. Importantly, AS1411 inhibited the growth of glioma xenograft and prolonged the survival time of glioma tumor-bearing mice. These results revealed a promising treatment of glioma by oligodeoxynucleotide aptamer.

  12. Induction of mitochondrial alternative oxidase in response to a cell signal pathway down-regulating the cytochrome pathway prevents programmed cell death.

    Science.gov (United States)

    Vanlerberghe, Greg C; Robson, Christine A; Yip, Justine Y H

    2002-08-01

    Treatment of tobacco (Nicotiana tabacum L. cv Petit Havana SR1) cells with cysteine (Cys) triggers a signal pathway culminating in a large loss of mitochondrial cytochrome (cyt) pathway capacity. This down-regulation of the cyt path likely requires events outside the mitochondrion and is effectively blocked by cantharidin or endothall, indicating that protein dephosphorylation is one critical process involved. Generation of reactive oxygen species, cytosolic protein synthesis, and Ca(2+) flux from organelles also appear to be involved. Accompanying the loss of cyt path is a large induction of alternative oxidase (AOX) protein and capacity. Induction of AOX allows the cells to maintain high rates of respiration, indicating that the lesion triggered by Cys is in the cyt path downstream of ubiquinone. Consistent with this, transgenic (AS8) cells unable to induce AOX (due to the presence of an antisense transgene) lose all respiratory capacity upon Cys treatment. This initiates in AS8 a programmed cell death pathway, as evidenced by the accumulation of oligonucleosomal fragments of DNA as the culture dies. Alternatively, wild-type cells remain viable and eventually recover their cyt path. Induction of AOX in response to a chemical inhibition of the cyt path (by antimycin A) is also dependent upon protein dephosphorylation and the generation of reactive oxygen species. Common events required for both down-regulation of the cyt path and induction of AOX may represent a mechanism to coordinate the biogenesis of these two electron transport paths. Such coordinate regulation may be necessary, not only to satisfy metabolic demands, but also to modulate the initiation of a programmed cell death pathway responsive to mitochondrial respiratory status.

  13. Interleukin-22 restored mitochondrial damage and impaired glucose-stimulated insulin secretion through down-regulation of uncoupling protein-2 in INS-1 cells.

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    Hu, Minling; Lin, Hanxiao; Yang, Li; Cheng, Yanzhen; Zhang, Hua

    2017-01-07

    Defective glucose-stimulated insulin secretion (GSIS) induced by chronic exposure to fatty acids is a hallmark of type 2 diabetes (T2D). Interleukin-22 (IL-22) has been shown to exert beneficial effects on insulin secretion and to protect pancreatic β-cells from stress. Moreover, uncoupling protein-2 (UCP-2) plays a central role in the regulation of GSIS and β-cell dysfunction, whereas the role of UCP-2 in IL-22-enhanced glycemic control under conditions of lipotoxicity remains unclear. In this present study, we investigated the effects of IL-22 on rat insulin-secreting cells (INS-1 cells) and the mechanisms that underlie IL-22 and lipotoxicity-impaired GSIS in vitro. Chronic palmitate (PA) treatment impaired insulin secretion and activated UCP-2 expression in INS-1 cells. Furthermore, in INS-1 cells, both reduced mitochondrial membrane potential (ΔΨm) and impaired GSIS induced by PA treatment were effectively reversed by an inhibitor of UCP-2 (genipin). Additionally, compared with the PA-treated group, INS-1 cells treated with IL-22 down-regulated UCP-2 expression, increased mitochondrial membrane potential, and restored GSIS. Together, our findings indicate that chronic exposure to PA could activate UCP-2, resulting in mitochondrial damage and impaired GSIS in INS-1 cells. We also suggest that IL-22 plays a protective role in this process via the down-regulation of UCP-2.

  14. Down-regulation of BLIMP1α by the EBV oncogene, LMP-1, disrupts the plasma cell differentiation program and prevents viral replication in B cells: implications for the pathogenesis of EBV-associated B-cell lymphomas.

    Science.gov (United States)

    Vrzalikova, Katerina; Vockerodt, Martina; Leonard, Sarah; Bell, Andrew; Wei, Wenbin; Schrader, Alexandra; Wright, Kenneth L; Kube, Dieter; Rowe, Martin; Woodman, Ciaran B; Murray, Paul G

    2011-06-02

    An important pathogenic event in Epstein-Barr virus (EBV)-associated lymphomas is the suppression of virus replication, which would otherwise lead to cell death. Because virus replication in B cells is intimately linked to their differentiation toward plasma cells, we asked whether the physiologic signals that drive normal B-cell differentiation are absent in EBV-transformed cells. We focused on BLIMP1α, a transcription factor that is required for plasma cell differentiation and that is inactivated in diffuse large B-cell lymphomas. We show that BLIMP1α expression is down-regulated after EBV infection of primary germinal center B cells and that the EBV oncogene, latent membrane protein-1 (LMP-1), is alone capable of inducing this down-regulation in these cells. Furthermore, the down-regulation of BLIMP1α by LMP-1 was accompanied by a partial disruption of the BLIMP1α transcriptional program, including the aberrant induction of MYC, the repression of which is required for terminal differentiation. Finally, we show that the ectopic expression of BLIMP1α in EBV-transformed cells can induce the viral lytic cycle. Our results suggest that LMP-1 expression in progenitor germinal center B cells could contribute to the pathogenesis of EBV-associated lymphomas by down-regulating BLIMP1α, in turn preventing plasma cell differentiation and induction of the viral lytic cycle.

  15. DMBT1 expression is down-regulated in breast cancer

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    Coggi G

    2004-08-01

    Full Text Available Abstract Background We studied the expression of DMBT1 (deleted in malignant brain tumor 1, a putative tumor suppressor gene, in normal, proliferative, and malignant breast epithelium and its possible relation to cell cycle. Methods Sections from 17 benign lesions and 55 carcinomas were immunostained with anti DMBT1 antibody (DMBTh12 and sections from 36 samples, were double-stained also with anti MCM5, one of the 6 pre-replicative complex proteins with cell proliferation-licensing functions. DMBT1 gene expression at mRNA level was assessed by RT-PCR in frozen tissues samples from 39 patients. Results Normal glands and hyperplastic epithelium in benign lesions displayed a luminal polarized DMBTh12 immunoreactivity. Normal and hyperplastic epithelium adjacent to carcinomas showed a loss of polarization, with immunostaining present in basal and perinuclear cytoplasmic compartments. DMBT1 protein expression was down-regulated in the cancerous lesions compared to the normal and/or hyperplastic epithelium adjacent to carcinomas (3/55 positive carcinomas versus 33/42 positive normal/hyperplastic epithelia; p = 0.0001. In 72% of cases RT-PCR confirmed immunohistochemical results. Most of normal and hyperplastic mammary cells positive with DMBTh12 were also MCM5-positive. Conclusions The redistribution and up-regulation of DMBT1 in normal and hyperplastic tissues flanking malignant tumours and its down-regulation in carcinomas suggests a potential role in breast cancer. Moreover, the concomitant expression of DMTB1 and MCM5 suggests its possible association with the cell-cycle regulation.

  16. Down-regulation of AP-4 inhibits proliferation, induces cell cycle arrest and promotes apoptosis in human gastric cancer cells.

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    Xinghua Liu

    Full Text Available BACKGROUND: AP-4 belongs to the basic helix-loop-helix leucine-zipper subgroup; it controls target gene expression, regulates growth, development and cell apoptosis and has been implicated in tumorigenesis. Our previous studies indicated that AP-4 was frequently overexpressed in gastric cancers and may be associated with the poor prognosis. The purpose of this study is to examine whether silencing of AP-4 can alter biological characteristics of gastric cancer cells. METHODS: Two specific siRNAs targeting AP-4 were designed, synthesized, and transfected into gastric cancer cell lines and human normal mucosa cells. AP-4 expression was measured with real-time quantitative PCR and Western blot. Cell proliferation and chemo-sensitivity were detected by CCK-8 assay. Cell cycle assay and apoptosis assay were performed by flow cytometer, and relative expression of cell cycle regulators were detected by real-time quantitative PCR and Western blot, expression of the factors involved in the apoptosis pathway were examined in mRNA and protein level. RESULTS: The expression of AP-4 was silenced by the siRNAs transfection and the effects of AP-4 knockdown lasted 24 to 96 hrs. The siRNA-mediated silencing of AP-4 suppressed the cellular proliferation, induced apoptosis and sensitized cancer cells to anticancer drugs. In addition, the expression level of p21, p53 and Caspase-9 were increased when AP-4 was knockdown, but the expression of cyclin D1, Bcl-2 and Bcl-x(L was inhibited. It didn't induce cell cycle arrest when AP-4 was knockdown in p53 defect gastric cancer cell line Kato-III. CONCLUSIONS: These results illustrated that gene silencing of AP-4 can efficiently inhibited cell proliferation, triggered apoptosis and sensitized cancer cells to anticancer drugs in vitro, suggesting that AP-4 siRNAs mediated silencing has a potential value in the treatment of human gastric cancer.

  17. Down-regulation of Yes Associated Protein 1 expression reduces cell proliferation and clonogenicity of pancreatic cancer cells.

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    Caroline H Diep

    Full Text Available BACKGROUND: The Hippo pathway regulates organ size by inhibiting cell proliferation and promoting cell apoptosis upon its activation. The Yes Associated Protein 1 (YAP1 is a nuclear effector of the Hippo pathway that promotes cell growth as a transcription co-activator. In human cancer, the YAP1 gene was reported as amplified and over-expressed in several tumor types. METHODS: Immunohistochemical staining of YAP1 protein was used to assess the expression of YAP1 in pancreatic tumor tissues. siRNA oligonucleotides were used to knockdown the expression of YAP1 and their effects on pancreatic cancer cells were investigated using cell proliferation, apoptosis, and anchorage-independent growth assays. The Wilcoxon signed-rank, Pearson correlation coefficient, Kendall's Tau, Spearman's Rho, and an independent two-sample t (two-tailed test were used to determine the statistical significance of the data. RESULTS: Immunohistochemistry studies in pancreatic tumor tissues revealed YAP1 staining intensities were moderate to strong in the nucleus and cytoplasm of the tumor cells, whereas the adjacent normal epithelial showed negative to weak staining. In cultured cells, YAP1 expression and localization was modulated by cell density. YAP1 total protein expression increased in the nuclear fractions in BxPC-3 and PANC-1, while it declined in HPDE6 as cell density increased. Additionally, treatment of pancreatic cancer cell lines, BxPC-3 and PANC-1, with YAP1-targeting siRNA oligonucleotides significantly reduced their proliferation in vitro. Furthermore, treatment with YAP1 siRNA oligonucleotides diminished the anchorage-independent growth on soft agar of pancreatic cancer cells, suggesting a role of YAP1 in pancreatic cancer tumorigenesis. CONCLUSIONS: YAP1 is overexpressed in pancreatic cancer tissues and potentially plays an important role in the clonogenicity and growth of pancreatic cancer cells.

  18. Down-regulation of phosphoglucomutase 3 mediates sulforaphane-induced cell death in LNCaP prostate cancer cells

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    Lee Hyo-Jeong

    2010-12-01

    Full Text Available Abstract Background Sulforaphane (SFN is an isothiocyanate found in cruciferous vegetables that exerts anti-oxidant, anti-inflammatory, anti-cancer and radio-sensitizing activities. Nonetheless, the mechanism responsible for SFN-induced cell death is not fully understood. In the present study, anti-cancer mechanism of SFN was elucidated in LNCaP prostate cancer cells. Results SFN exerted cytotoxicity and increased TUNEL positive cells in a concentration-dependent manner in LNCaP cells. Proteomics study revealed that levels of nine proteins including tubulin β-2, phosphoglucomutase-3 (PGM3, melanoma-derived leucine zipper containing extra-nuclear factor, activin A type I receptor precursor, smoothelin-A, KIA0073, hypothetical protein LOC57691 and two unnamed proteins were changed over 8 folds in SFN treated LNCaP cells compared to untreated control. We have further confirmed that SFN reduced PGM3 expression with western blotting and showed that PGM3 siRNA enhanced cytotoxicity demonstrated by cell morphology and TUNEL assays in LNCaP cells. Conclusion Taken together, these findings suggest that PGM3 plays a role in mediating SFN-induced cell death in LNCaP cells, and is a potential molecular therapeutic target for prostate cancer.

  19. pH-responsive artemisinin derivatives and lipid nanoparticle formulations inhibit growth of breast cancer cells in vitro and induce down-regulation of HER family members.

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    Yitong J Zhang

    Full Text Available Artemisinin (ART dimers show potent anti-proliferative activities against breast cancer cells. To facilitate their clinical development, novel pH-responsive artemisinin dimers were synthesized for liposomal nanoparticle formulations. A new ART dimer was designed to become increasingly water-soluble as pH declines. The new artemisinin dimer piperazine derivatives (ADPs remained tightly associated with liposomal nanoparticles (NPs at neutral pH but were efficiently released at acidic pH's that are known to exist within solid tumors and organelles such as endosomes and lysosomes. ADPs incorporated into nanoparticles down regulated the anti-apoptotic protein, survivin, and cyclin D1 when incubated at low concentrations with breast cancer cell lines. We demonstrate for the first time, for any ART derivative, that ADP NPs can down regulate the oncogenic protein HER2, and its counterpart, HER3 in a HER2+ cell line. We also show that the wild type epidermal growth factor receptor (EGFR or HER1 declines in a triple negative breast cancer (TNBC cell line in response to ADP NPs. The declines in these proteins are achieved at concentrations of NP109 at or below 1 µM. Furthermore, the new artemisinin derivatives showed improved cell-proliferation inhibition effects compared to known dimer derivatives.

  20. HACE1 is a putative tumor suppressor gene in B-cell lymphomagenesis and is down-regulated by both deletion and epigenetic alterations.

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    Bouzelfen, Abdelilah; Alcantara, Marion; Kora, Hafid; Picquenot, Jean-Michel; Bertrand, Philippe; Cornic, Marie; Mareschal, Sylvain; Bohers, Elodie; Maingonnat, Catherine; Ruminy, Philippe; Adriouch, Sahil; Boyer, Olivier; Dubois, Sydney; Bastard, Christian; Tilly, Hervé; Latouche, Jean-Baptiste; Jardin, Fabrice

    2016-06-01

    HECT domain and ankyrin repeat containing E3 ubiquitin protein ligase 1, HACE1, located on chromosome 6q, encodes an E3 ubiquitin ligase and is downregulated in many human tumors. Here, we report HACE1 as a candidate tumor suppressor gene down-regulated by a combination of deletion and epigenetic mechanisms. HACE1 deletions were observed in 40% of B-cell lymphoma tumors. Hypermethylation of the HACE1 promoter CpG177 island was found in 60% (68/111) of cases and in all tested B-cell lymphoma lines. Using HDAC inhibitors, we observed predominantly inactive chromatin conformation (methylated H3 histones H3K9me2) in HACE1 gene promoter region. We demonstrated in Ramos and Raji cells that down-regulation of HACE1 expression was associated with a significant decrease in apoptosis and an accumulation of cells in the S and G2/M phases. Our experiments indicate that HACE1 can act as a haploinsufficient tumor suppressor gene in most B-cell lymphomas and can be downregulated by deacetylation of its promoter region chromatin, which makes HACE1 a potential target for HDAC inhibitors.

  1. Selective oral ROCK2 inhibitor down-regulates IL-21 and IL-17 secretion in human T cells via STAT3-dependent mechanism.

    Science.gov (United States)

    Zanin-Zhorov, Alexandra; Weiss, Jonathan M; Nyuydzefe, Melanie S; Chen, Wei; Scher, Jose U; Mo, Rigen; Depoil, David; Rao, Nishta; Liu, Ben; Wei, Jianlu; Lucas, Sarah; Koslow, Matthew; Roche, Maria; Schueller, Olivier; Weiss, Sara; Poyurovsky, Masha V; Tonra, James; Hippen, Keli L; Dustin, Michael L; Blazar, Bruce R; Liu, Chuan-ju; Waksal, Samuel D

    2014-11-25

    Rho-associated kinase 2 (ROCK2) regulates the secretion of proinflammatory cytokines and the development of autoimmunity in mice. Data from a phase 1 clinical trial demonstrate that oral administration of KD025, a selective ROCK2 inhibitor, to healthy human subjects down-regulates the ability of T cells to secrete IL-21 and IL-17 by 90% and 60%, respectively, but not IFN-γ in response to T-cell receptor stimulation in vitro. Pharmacological inhibition with KD025 or siRNA-mediated inhibition of ROCK2, but not ROCK1, significantly diminished STAT3 phosphorylation and binding to IL-17 and IL-21 promoters and reduced IFN regulatory factor 4 and nuclear hormone RAR-related orphan receptor γt protein levels in T cells derived from healthy subjects or rheumatoid arthritis patients. Simultaneously, treatment with KD025 also promotes the suppressive function of regulatory T cells through up-regulation of STAT5 phosphorylation and positive regulation of forkhead box p3 expression. The administration of KD025 in vivo down-regulates the progression of collagen-induced arthritis in mice via targeting of the Th17-mediated pathway. Thus, ROCK2 signaling appears to be instrumental in regulating the balance between proinflammatory and regulatory T-cell subsets. Targeting of ROCK2 in man may therefore restore disrupted immune homeostasis and have a role in the treatment of autoimmunity.

  2. Autophagy down regulates pro-inflammatory mediators in BV2 microglial cells and rescues both LPS and alpha-synuclein induced neuronal cell death

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    Bussi, Claudio; Ramos, Javier Maria Peralta; Arroyo, Daniela S.; Gaviglio, Emilia A.; Gallea, Jose Ignacio; Wang, Ji Ming; Celej, Maria Soledad; Iribarren, Pablo

    2017-01-01

    Autophagy is a fundamental cellular homeostatic mechanism, whereby cells autodigest parts of their cytoplasm for removal or turnover. Neurodegenerative disorders are associated with autophagy dysregulation, and drugs modulating autophagy have been successful in several animal models. Microglial cells are phagocytes in the central nervous system (CNS) that become activated in pathological conditions and determine the fate of other neural cells. Here, we studied the effects of autophagy on the production of pro-inflammatory molecules in microglial cells and their effects on neuronal cells. We observed that both trehalose and rapamycin activate autophagy in BV2 microglial cells and down-regulate the production of pro-inflammatory cytokines and nitric oxide (NO), in response to LPS and alpha-synuclein. Autophagy also modulated the phosphorylation of p38 and ERK1/2 MAPKs in BV2 cells, which was required for NO production. These actions of autophagy modified the impact of microglial activation on neuronal cells, leading to suppression of neurotoxicity. Our results demonstrate a novel role for autophagy in the regulation of microglial cell activation and pro-inflammatory molecule secretion, which may be important for the control of inflammatory responses in the CNS and neurotoxicity. PMID:28256519

  3. Transmembrane neural cell-adhesion molecule (NCAM), but not glycosyl-phosphatidylinositol-anchored NCAM, down-regulates secretion of matrix metalloproteinases

    DEFF Research Database (Denmark)

    Edvardsen, K; Chen, W; Rucklidge, G;

    1993-01-01

    During embryogenesis interactions between cells and extracellular matrix play a central role in the modulation of cell motility, growth, and differentiation. Modulation of matrix structure is therefore crucial during development; extracellular matrix ligands, their receptors, extracellular...... proteinases, and proteinase inhibitors all participate in the construction, maintenance, and remodeling of extracellular matrix by cells. The neural cell-adhesion molecule (NCAM)-negative rat glioma cell line BT4Cn secretes substantial amounts of metalloproteinases, as compared with its NCAM-positive mother...... cell line BT4C. We have transfected the BT4Cn cell line with cDNAs encoding the human NCAM-B and -C isoforms. We report here that the expression of transmembrane NCAM-B, but not of glycosyl-phosphatidylinositol-linked NCAM-C, induces a down-regulation of 92-kDa gelatinase (matrix metalloproteinase 9...

  4. Effects of RNA interference-induced tryptase down-regulation in P815 cells on IL-6 and TNF-α release of endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Yi-feng JIANG; Feng-di ZHAO; Xiao-bo LI; Yan-xia NING; Xiu-ling ZHI; Rui-zhe QIAN; Lian-hua YIN

    2008-01-01

    Objective:To explore the effects of down-regulated tryptase expression in mast cells on the synthesis and release of interleukin-6(IL-6)and tumor necrosis factor-alpha(TNF-α) of vascular endothelial cells.Methods:Tryptase-siRNA (small-interfering RNA)vector was constructed to inhibit tryptase expression in P815 cells.The medium of P815 cells treated by the tryptase-siRNA(RNAi-P815 group)or pure vector(P815 group)was collected and used to culture bEnd.3 cells.The messenger RNAs (mRNAs)of IL-6 and TNF-a in bEnd.3 cells and their protein levels in the medium were measured by reverse transcription polymerase chain reaction(RT-PCR)and enzyme-linked immunosorbent assay(ELISA),respectively.Results:IL-6 and TNF-α mRNAs in bEnd.3 cells cultured in RNAi-P815-conditioned medium decreased significantly compared to those in P815-conditioned medium.Consistently.IL-6 and TNF-α protein levels in the medium of bEnd.3 of RNAi-P815 group were lower than those of P815 group.Conclusion:Reduced tryptase expression significantly inhibited the synthesis and release of IL-6 and TNF-α in vascular endothelial cells.RNA interference targeting tryptasc expression may be a new anti-inflammatory strategy for vascular diseases.

  5. HIV infection enhances TRAIL-induced cell death in macrophage by down-regulating decoy receptor expression and generation of reactive oxygen species.

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    Dan-Ming Zhu

    Full Text Available BACKGROUND: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL could induce apoptosis of HIV-1-infected monocyte-derived macrophage (MDM, but the molecular mechanisms are not well understood. METHODOLOGY/PRINCIPAL FINDINGS: By using an HIV-1 Env-pseudotyped virus (HIV-1 PV-infected MDM cell model we demonstrate that HIV-1 PV infection down-regulates the expression of TRAIL decoy receptor 1 (DcR1 and 2 (DcR2, and cellular FLICE-inhibitory protein (c-FLIP, but dose not affect the expression of death receptor 4 and 5 (DR4, DR5, and Bcl-2 family members in MDM cells. Furthermore, recombinant soluble TRAIL and an agonistic anti-DR5 antibody, AD5-10, treatment stimulates reactive oxygen species (ROS generation and JNK phosphorylation. CONCLUSIONS/SIGNIFICANCE: HIV infection facilitates TRIAL-induced cell death in MDM by down-regulating the expression of TRAIL decoy receptors and intracellular c-FLIP. Meanwhile, the agonistic anti-DR5 antibody, AD5-10, induces apoptosis synergistically with TRAIL in HIV-1-infected cells. ROS generation and JNK phosphorylation are involved in this process. These findings potentiate clinical usage of the combination of TRAIL and AD5-10 in eradication of HIV-infected macrophage and AIDS.

  6. Suppression of c-Myc is involved in multi-walled carbon nanotubes' down-regulation of ATP-binding cassette transporters in human colon adenocarcinoma cells.

    Science.gov (United States)

    Wang, Zhaojing; Xu, Yonghong; Meng, Xiangning; Watari, Fumio; Liu, Hudan; Chen, Xiao

    2015-01-01

    Over-expression of ATP-binding cassette (ABC) transporters, a large family of integral membrane proteins that decrease cellular drug uptake and accumulation by active extrusion, is one of the major causes of cancer multi-drug resistance (MDR) that frequently leads to failure of chemotherapy. Carbon nanotubes (CNTs)-based drug delivery devices hold great promise in enhancing the efficacy of cancer chemotherapy. However, CNTs' effects on the ABC transporters remain under-investigated. In this study, we found that multiwalled carbon nanotubes (MWCNTs) reduced transport activity and expression of ABC transporters including ABCB1/Pgp and ABCC4/MRP4 in human colon adenocarcinoma Caco-2 cells. Proto-oncogene c-Myc, which directly regulates ABC gene expression, was concurrently decreased in MWCNT-treated cells and forced over-expression of c-Myc reversed MWCNTs' inhibitory effects on ABCB1 and ABCC4 expression. MWCNT-cell membrane interaction and cell membrane oxidative damage were observed. However, antioxidants such as vitamin C, β-mecaptoethanol and dimethylthiourea failed to antagonize MWCNTs' down-regulation of ABC transporters. These data suggest that MWCNTs may act on c-Myc, but not through oxidative stress, to down-regulate ABC transporter expression. Our findings thus shed light on CNTs' novel cellular effects that may be utilized to develop CNTs-based drug delivery devices to overcome ABC transporter-mediated cancer chemoresistance.

  7. Down-regulation of wt1 expression in leukemia cell lines as part of apoptotic effect in arsenic treatment using two compounds.

    Science.gov (United States)

    Glienke, Wolfgang; Chow, Kai U; Bauer, Nina; Bergmann, Lothar

    2006-08-01

    Arsenic trioxide (As2O3) induces remission in patients with acute promyelocytic leukemia (APL). To better understand molecular mechanisms of arsenic actions, this study investigated the effect of two different arsenic compounds on gene expression of apoptosis and cellular proliferation related genes. The Wilms' tumor gene (wt1) is up-regulated in acute myeloid leukemia (AML) and a variety of leukemia cell lines. The expression of wt1 in these cells is proposed to have an anti-apoptotic effect. HL-60 and K562 were treated with arsenic trioxide (As2O3) and sodium arsenite (NaAsO2) at concentrations between 0 - 10 microM for up to 48 h. The induction of apoptosis was accompanied by down-regulation of hTERT and wt1 mRNA and protein expression but up-regulation of par-4. Low concentrations of 0.1 microM arsenic induced expression of the anti-apoptotic bcl-2 gene in both cell lines HL-60 and K562. There were no major differences encountered between compounds. After arsenic treatment of the leukemia cell lines HL-60 and K562 the up-regulation of par-4 may contribute to the induction of apoptosis rather than down-regulation of bcl-2. The therapeutic effect of arsenic is the induction of apoptosis by modulating the gene expression profile of pro- and anti-apoptotic genes including the wt1 gene.

  8. In vitro mechanism for down-regulation of ERalpha expression by epigallocatechin gallate in ER+/PR+ human breast cancer cells

    Science.gov (United States)

    De Amicis, Francesca; Russo, Alessandra; Avena, Paola; Santoro, Marta; Vivacqua, Adele; Bonofiglio, Daniela; Mauro, Loredana; Aquila, Saveria; Tramontano, Donatella; Fuqua, Suzanne AW; Andò, Sebastiano

    2015-01-01

    Scope Exposure of the breast to estrogens and other sex hormones is an important cancer risk factor and estrogen receptor down-regulators are attracting significant clinical interest. Epigallocatechin gallate (EGCG), a polyphenolic compound found in green tea, has gained considerable attention for its antitumor properties. Here we aimed to investigate the molecular mechanisms through which EGCG regulates ERα expression in ER+ PR+ breast cancer cells. Material and Methods Western blotting analysis, real time PCR and transient transfections of deletion fragments of the ERα gene promoter show that EGCG down-regulates ERα protein, mRNA and gene promoter activity with a concomitant reduction of ERα genomic and non genomic signal. These events occur through p38MAPK/CK2 activation, causing the release from Hsp90 of PR-B and its consequent nuclear translocation as evidenced by immunofluorescence studies. EMSA and ChIP assay reveal that, upon EGCG treatment, PR-B is recruited at the half PRE site on ERα promoter. This is concomitant with the formation of a corepressor complex containing NCoR and HDAC1 while RNA polymerase II is displaced. The events are crucially mediated by PR-B isoform, since they are abrogated with PR-B siRNA. Conclusions Our data provide evidence for a mechanism by which EGCG down-regulates ERα and explain the inhibitory action of EGCG on the proliferation of ER+ PR+ cancer cells tested. We suggest that the EGCG/PR-B signaling should be further exploited for clinical approach. PMID:23322423

  9. CHCHD2 is down-regulated in neuronal cells differentiated from iPS cells derived from patients with lissencephaly.

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    Shimojima, Keiko; Okumura, Akihisa; Hayashi, Masaharu; Kondo, Takayuki; Inoue, Haruhisa; Yamamoto, Toshiyuki

    2015-10-01

    The human cerebral cortex is peculiar for a six-layered cellular-sheet structure with convolution, which is a consequence of neuronal migration. Dysfunctions of the pathways contributing to this mechanism typically lead to lissencephaly manifesting smooth brain surfaces. To investigate the unknown mechanism underlying neuronal migration disorders, we generated induced pluripotent stem (iPS) cells from two patients with lissencephaly. Whole gene expression study for iPS cells derived from a patient with a LIS1 deletion showed reduced expression of the coiled-coil-helix-coiled-coil-helix domain containing 2 gene (CHCHD2), which was also confirmed in iPS cells derived from a patient with a TUBA1A mutation. CHCHD2 expression was detected in neuronal cells differentiated from normal iPS cells in a time-dependent manner, as well as in the brain of a fetus at 26-28 week gestational age, suggesting development-dependent expression. Migrating neuronal cells showed CHCHD2 expression, suggesting its functional relevance to neuronal migration.

  10. Fucoidan induces G1 arrest of the cell cycle in EJ human bladder cancer cells through down-regulation of pRB phosphorylation

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    Hye Young Park

    2015-06-01

    Full Text Available AbstractFucoidan, a sulfated polysaccharide found in marine algae and brown seaweeds, has been shown to inhibit the in vitro growth of human cancer cells. This study was conducted in cultured human bladder cancer EJ cells to elucidate the possible mechanisms by which fucoidan exerts its anti-proliferative activity, which until now has remained poorly understood. Fucoidan treatment of EJ cells resulted in dose-dependent inhibition of cell growth and induced apoptotic cell death. Flow cytometric analysis revealed that fucoidan led to G1 arrest in cell cycle progression. It was associated with down-regulation of cyclin D1, cyclin E, and cyclin-dependent-kinases (Cdks in a concentration-dependent manner, without any change in Cdk inhibitors, such as p21 and p27. Furthermore, dephosphorylation of retinoblastoma protein (pRB by this compound was associated with enhanced binding of pRB with the transcription factors E2F-1 and E2F-4. Overall, our results demonstrate that fucoidan possesses anticancer activity potential against bladder cancer cells by inhibiting pRB phosphorylation.

  11. miR-338-3p Is Down-Regulated by Hepatitis B Virus X and Inhibits Cell Proliferation by Targeting the 3′-UTR Region of CyclinD1

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    Xiaoyu Fu

    2012-07-01

    Full Text Available Hepatitis B virus X protein (HBx is recognized as an oncogene in hepatocellular carcinoma (HCC. HBx regulates microRNA expression, including down-regulating miR-338-3p in LO2 cells. Here, we investigated miR-338-3p function in HBx-mediated hepatocarcinogenesis. In 23 HBV-infected HCC clinical patient tumor and adjacent non-tumor control tissues, 17 and 19 tumors expressed HBx mRNA and protein, respectively. When considered as a group, HBV-infected HCC tumors had lower miR-338-3p expression than controls; however, miR-338-3p was only significantly down-regulated in HBx-positive tumors, indicating that HBx inversely correlated with miR-338-3p. Functional characterization of miR-338-3p indicated that miR-338-3p mimics inhibited cell proliferation by inducing cell cycle arrest at the G1/S phase as assessed by EdU and cell cycle assays in HBx-expressing LO2 cells. CyclinD1, containing two putative miR-338-3p targets, was confirmed as a direct target using 3′-UTR luciferase reporter assays from cells transfected with mutated binding sites. Mutating the 2397–2403 nt binding site conferred the greatest resistance to miR-338-3p suppression of CyclinD1, indicating that miR-338-3p suppresses CyclinD1 at this site. Overall, this study demonstrates that miR-338-3p inhibits proliferation by regulating CyclinD1, and HBx down-regulates miR-338-3p in HCC. This newly identified miR-338-3p/CyclinD1 interaction provides novel insights into HBx-mediated hepatocarcinogenesis and may facilitate therapeutic development against HCC.

  12. Down-regulation of zinc transporter 8 (SLC30A8) in pancreatic beta-cells promotes cell survival.

    Science.gov (United States)

    The pancreatic islet contains high levels of zinc in granular vesicles of ß-cells where insulin is matured, crystallized, and stored before secretion. Zinc is an essential co-factor for insulin crystallization forming dense cores in secretory granules. In insulin-containing secretory granules, zinc ...

  13. Effects of stereochemistry, saturation, and hydrocarbon chain length on the ability of synthetic constrained azacyclic sphingolipids to trigger nutrient transporter down-regulation, vacuolation, and cell death.

    Science.gov (United States)

    Perryman, Michael S; Tessier, Jérémie; Wiher, Timothy; O'Donoghue, Heather; McCracken, Alison N; Kim, Seong M; Nguyen, Dean G; Simitian, Grigor S; Viana, Matheus; Rafelski, Susanne; Edinger, Aimee L; Hanessian, Stephen

    2016-09-15

    Constrained analogs containing a 2-hydroxymethylpyrrolidine core of the natural sphingolipids sphingosine, sphinganine, N,N-dimethylsphingosine and N-acetyl variants of sphingosine and sphinganine (C2-ceramide and dihydro-C2-ceramide) were synthesized and evaluated for their ability to down-regulate nutrient transporter proteins and trigger cytoplasmic vacuolation in mammalian cells. In cancer cells, the disruptions in intracellular trafficking produced by these sphingolipids lead to cancer cell death by starvation. Structure activity studies were conducted by varying the length of the hydrocarbon chain, the degree of unsaturation and the presence or absence of an aryl moiety on the appended chains, and stereochemistry at two stereogenic centers. In general, cytotoxicity was positively correlated with nutrient transporter down-regulation and vacuolation. This study was intended to identify structural and functional features in lead compounds that best contribute to potency, and to develop chemical biology tools that could be used to isolate the different protein targets responsible for nutrient transporter loss and cytoplasmic vacuolation. A molecule that produces maximal vacuolation and transporter loss is expected to have the maximal anti-cancer activity and would be a lead compound.

  14. Nerve growth factor blocks the glucose-induced down-regulation of caveolin-1 expression in Schwann cells via p75 neurotrophin receptor signaling.

    Science.gov (United States)

    Tan, Wenbin; Rouen, Shefali; Barkus, Kristin M; Dremina, Yelena S; Hui, Dongwei; Christianson, Julie A; Wright, Douglas E; Yoon, Sung Ok; Dobrowsky, Rick T

    2003-06-20

    Altered neurotrophism in diabetic peripheral neuropathy (DPN) is associated in part with substantial degenerative changes in Schwann cells (SCs) and an increased expression of the p75 neurotrophin receptor (p75NTR). Caveolin-1 (Cav-1) is highly expressed in adult SCs, and changes in its expression can regulate signaling through Erb B2, a co-receptor that mediates the effects of neuregulins in promoting SC growth and differentiation. We examined the hypothesis that hyperglycemia-induced changes in Cav-1 expression and p75NTR signaling may contribute to altered neurotrophism in DPN by modulating SC responses to neuregulins. In an animal model of type 1 diabetes, hyperglycemia induced a progressive decrease of Cav-1 in SCs of sciatic nerve that was reversed by insulin therapy. Treatment of primary neonatal SCs with 20-30 mm d-glucose, but not l-glucose, was sufficient to inhibit transcription from the Cav-1 promoter and decrease Cav-1 mRNA and protein expression. Hyperglycemia prolonged the kinetics of Erb B2 phosphorylation and significantly enhanced the mitogenic response of SCs to neuregulin1-beta1, and this effect was mimicked by the forced down-regulation of Cav-1. Intriguingly, nerve growth factor antagonized the enhanced mitogenic response of SCs to neuregulin1-beta1 and inhibited the glucose-induced down-regulation of Cav-1 transcription, mRNA, and protein expression through p75NTR-dependent activation of JNK. Our data suggest that Cav-1 down-regulation may contribute to altered neurotrophism in DPN by enhancing the response of SCs to neuregulins and that p75NTR-mediated JNK activation may provide a mechanism for the neurotrophic modulation of hyperglycemic stress.

  15. Collagen I-induced dendritic cells activation is regulated by TNF-alpha production through down-regulation of IRF4.

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    Poudel, Barun; Ki, Hyeon-Hui; Lee, Young-Mi; Kim, Dae-Ki

    2015-03-01

    Previously we have shown that collagen I enhances the maturation and function of dendritic cells (DCs). Inflammatory mediators such as tumour necrosis factor (TNF)- alpha, interleukin (IL)-1 beta and lipopolysaccharide (LPS) are also known to activate DCs. Here we investigated the involvement of TNF-alpha on the collagen I-induced DCs activation. TNF-a neutralization inhibited collagen I-induced IL-12 secretions by DCs. Additionally, we observed suppression of collagen I-induced costimulatory molecules expression along with down-regulation of genes involved in DCs activation pathway. Furthermore, TNF- alpha inhibition upon collagen Istimulation up-regulated the expression of interferon regulatory transcription factor IRF4, when compared to collagen I only treated cells. Collectively, our data demonstrate that collagen I induce TNF- alpha production, which is crucial for the activation and function of DCs, through down-regulation of IRF4, and implicates the importance in development of anti- TNF-alpha therapeutics for several inflammatory diseases.

  16. Down-regulation of Bmi-1 by RNA interference in Jurkat cells%RNAi抑制Bmi-1基因表达对Jurkat细胞的影响

    Institute of Scientific and Technical Information of China (English)

    Shangen Zheng; Qibin Jing; Yaqiong Zheng; Yinjuan Ding; Qianchuan Huang; Guoqiang Zhao

    2012-01-01

    Objective: The aim of our study was to investigate the effects of down-regulation Bmi-1 by RNA interference (RNAi) in T Lymphocytic leukemia Jurkat cells. Methods: Two complementary oligonucleotide strands were synthesized based on the siRNA sequence targeting Bmi-1 gene. After annealing, siRNA strands were recombined into the pRNATU6.2 vector, and then DNA sequencing was carried out following transformation and amplification. The recombinant was transfected into Jurkat cells with liposomes. Positive colonies were obtained through G418 selection. The mRNA and protein expressions of Bmi-1 were detected by RT-PCR and Western-blot, respectively. Effects of Bmi-1 silence on cell proliferation, cell cycle and cell aging of Jurkat cells were detected by MTT assay, flow cytometry, colony formation assay and SA-β-Gal staining, respectively. Results: The siRNA recombinant targeting Bmi-1 gene was successfully constructed. All three siRNA recombinants could significantly inhibit the expression of Bmi-1. The siRNA targeting 825nt-843nt (GACCAGACCACTACT GAAT) has the strongest inhibitory effect on Bmi-1 expression, with almost complete inhibition on Bmi-1 mRNA and protein expressions. Compared with the non-transfection group and the empty vector group, growth velocity and colony formation ability were significantly decreased, while the proportion of cells in G1 phase and the percentage of senile cells were significantly increased in highly transfected group (P < 0.05). Conclusion: Down-regulation Bmi-1 by RNA interference (RNAi) could significantly inhibit the growth of Jurkat cells in vitro.

  17. BAFF induces spleen CD4{sup +} T cell proliferation by down-regulating phosphorylation of FOXO3A and activates cyclin D2 and D3 expression

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    Ji, Fang; Chen, Rongjing [Department of Orthodontics, Ninth People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai (China); Liu, Baojun [Laboratory of Lung, Inflammation and Cancers, Huashan Hospital, Fudan University, Shanghai (China); Zhang, Xiaoping [Department of Nuclear Medicine, Shanghai 10th People' s Hospital, Tongji University School of Medicine, Shanghai 200072 (China); Han, Junli; Wang, Haining [Department of General Dentistry, Ninth People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai (China); Shen, Gang [Department of Orthodontics, Ninth People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai (China); Tao, Jiang, E-mail: taojiang2012@yahoo.cn [Department of General Dentistry, Ninth People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai (China)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer Firstly analyze the mechanism of BAFF and anti-CD3 co-stimulation on purified mouse splenic CD4{sup +} T cells. Black-Right-Pointing-Pointer Carrying out siRNA technology to study FOXO3A protein function. Black-Right-Pointing-Pointer Helpful to understand the T cell especially CD4{sup +} T cell's role in immunological reaction. -- Abstract: The TNF ligand family member 'B cell-activating factor belonging to the TNF family' (BAFF, also called BLyS, TALL-1, zTNF-4, and THANK) is an important survival factor for B and T cells. In this study, we show that BAFF is able to induce CD4{sup +} spleen T cell proliferation when co-stimulated with anti-CD3. Expression of phosphorylated FOXO3A was notably down-regulated and cyclins D2 and D3 were up-regulated and higher in the CD4{sup +} T cells when treated with BAFF and anti-CD3, as assessed by Western blotting. Furthermore, after FOXO3A was knocked down, expression of cyclin D1 was unchanged, compared with control group levels, but the expression of cyclins D2 and D3 increased, compared with the control group. In conclusion, our results suggest that BAFF induced CD4{sup +} spleen T cell proliferation by down-regulating the phosphorylation of FOXO3A and then activating cyclin D2 and D3 expression, leading to CD4{sup +} T cell proliferation.

  18. Down-regulation of human leukocyte antigens class I on peripheral T lymphocytes and NK cells from subjects in region of high-incidence gastrointestinal tumor

    Institute of Scientific and Technical Information of China (English)

    ZHANG Zhi-mian; LI Ying-jie; GUAN Xiao; YANG Xiao-yun; GAO Xi-mei; YANG Xiao-jing; WANG Li-shui; ZOU Xiong

    2011-01-01

    Background Many types of human tumors can suppress the immune system to enhance their survival. Loss or down-regulation of human leukocyte antigens (HLA) class I on tumors is considered to be a major mechanism of tumor immune escape. Our previous studies found that HLA class I on peripheral-blood mononuclear cells was significantly lower in gastric cancer patients. The present study made an analysis of HLA class I expression on peripheral-blood T lymphocytes and NK cells from subjects of Lijiadian village, a village with high-incidence gastrointestinal tumor. Methods A total of 181 villagers from Lijiadian village and 153 normal controls from the Department of Health Examination Center were enrolled in this study. Using a multi-tumor markers detection system, these villagers were divided into two groups: high-risk group (tumor markers positive group) and low-risk group (tumor markers negative group). The percentage of T lymphocytes and NK cells and levels of HLA class I on their surface were determined in these subjects by flow cytometry.Results Percentages of T lymphocytes and NK cells in peripheral-blood mononuclear cells did not vary with age. The expression level of HLA class I on peripheral T lymphocytes and NK cells was not affected by age or gender, but was significantly down-regulated in Lijiadian villagers (P<0.05), especially on the surface of NK cells (P<0.01). Compared with the low-risk group, there was a significant reduction of HLA class I on peripheral T lymphocytes (P <0.05) and NK cells (P <0.05) in the high-risk group.Conclusions HLA class I on peripheral T lymphocytes and NK cells may be involved in tumorigenesis and development of gastrointestinal tumor, and understanding their changes in expression may provide new insights into the mechanism of tumor immunity.

  19. Melatonin down-regulates hTERT expression induced by either natural estrogens (17beta-estradiol) or metalloestrogens (cadmium) in MCF-7 human breast cancer cells.

    Science.gov (United States)

    Martínez-Campa, Carlos M; Alonso-González, Carolina; Mediavilla, Maria D; Cos, Samuel; González, Alicia; Sanchez-Barcelo, Emilio J

    2008-09-18

    The goal was to evaluate whether melatonin (Mel) down-regulates hTERT expression induced by 17beta-estradiol (E(2)) or cadmium (Cd) in breast cancer cells. We found that: (a) Mel inhibits E(2) or Cd-induced hTERT transcription in hTERT-Luc transfected MCF-7 cells, (b) Mel significantly reduces E(2)- and Cd-mediated hTERT transactivation triggered by ERalpha in transfected HeLa cells, (c) Mel inhibits hTERT expression induced by E(2) or Cd in MCF-7 cells. Melatonin inhibition of telomerase activity supports a possible role in treatment of estrogen-dependent tumors or carcinogenesis by environmental or occupational exposure to xenoestrogens.

  20. Clitocine reversal of P-glycoprotein associated multi-drug resistance through down-regulation of transcription factor NF-κB in R-HepG2 cell line.

    Directory of Open Access Journals (Sweden)

    Jianguo Sun

    Full Text Available Multidrug resistance (MDR is one of the major reasons for failure in cancer chemotherapy and its suppression may increase the efficacy of therapy. The human multidrug resistance 1 (MDR1 gene encodes the plasma membrane P-glycoprotein (P-gp that pumps various anti-cancer agents out of the cancer cell. R-HepG2 and MES-SA/Dx5 cells are doxorubicin induced P-gp over-expressed MDR sublines of human hepatocellular carcinoma HepG2 cells and human uterine carcinoma MES-SA cells respectively. Herein, we observed that clitocine, a natural compound extracted from Leucopaxillus giganteus, presented similar cytotoxicity in multidrug resistant cell lines compared with their parental cell lines and significantly suppressed the expression of P-gp in R-HepG2 and MES-SA/Dx5 cells. Further study showed that the clitocine increased the sensitivity and intracellular accumulation of doxorubicin in R-HepG2 cells accompanying down-regulated MDR1 mRNA level and promoter activity, indicating the reversal effect of MDR by clitocine. A 5'-serial truncation analysis of the MDR1 promoter defined a region from position -450 to -193 to be critical for clitocine suppression of MDR1. Mutation of a consensus NF-κB binding site in the defined region and overexpression of NF-κB p65 could offset the suppression effect of clitocine on MDR1 promoter. By immunohistochemistry, clitocine was confirmed to suppress the protein levels of both P-gp and NF-κB p65 in R-HepG2 cells and tumors. Clitocine also inhibited the expression of NF-κB p65 in MES-SA/Dx5. More importantly, clitocine could suppress the NF-κB activation even in presence of doxorubicin. Taken together; our results suggested that clitocine could reverse P-gp associated MDR via down-regulation of NF-κB.

  1. Moraxella catarrhalis decreases antiviral innate immune responses by down-regulation of TLR3 via inhibition of p53 in human bronchial epithelial cells.

    Science.gov (United States)

    Heinrich, Annina; Haarmann, Helge; Zahradnik, Sabrina; Frenzel, Katrin; Schreiber, Frauke; Klassert, Tilman E; Heyl, Kerstin A; Endres, Anne-Sophie; Schmidtke, Michaela; Hofmann, Jörg; Slevogt, Hortense

    2016-06-01

    Chronic obstructive pulmonary disease (COPD) is complicated by infectious exacerbations with acute worsening of respiratory symptoms. Coinfections of bacterial and viral pathogens are associated with more severe exacerbations. Moraxella catarrhalis is one of the most frequent lower respiratory tract pathogens detected in COPD. We therefore studied the impact of M. catarrhalis on the antiviral innate immune response that is mediated via TLR3 and p53. Molecular interactions between M. catarrhalis and normal human bronchial epithelial (NHBE) cells as well as Beas-2B cells were studied using flow cytometry, quantitative PCR analysis, chromatin immunoprecipitation, RNA interference, and ELISA. M. catarrhalis induces a significant down-regulation of TLR3 in human bronchial epithelial cells. In M. catarrhalis-infected cells, expression of p53 was decreased. We detected a reduced binding of p53 to the tlr3 promoter, resulting in reduced TLR3 gene transcription. M. catarrhalis diminished the TLR3-dependent secretion of IFN-β, IFN-λ, and chemokine (C-X-C motif) ligand 8. In addition in M. catarrhalis infected cells, expression of rhinovirus type 1A RNA was increased compared with uninfected cells. M. catarrhalis reduces antiviral defense functions of bronchial epithelial cells, which may increase susceptibility to viral infections.-Heinrich, A., Haarmann, H., Zahradnik, S., Frenzel, K., Schreiber, F., Klassert, T. E., Heyl, K. A., Endres, A.-S., Schmidtke, M., Hofmann, J., Slevogt, H. Moraxella catarrhalis decreases antiviral innate immune responses by down-regulation of TLR3 via inhibition of p53 in human bronchial epithelial cells.

  2. Down regulation of the TCR complex CD3 ζ-chain on CD3+ T cells: a potential mechanism for helminth mediated immune modulation

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    Laura Jane Appleby

    2015-02-01

    Full Text Available The CD3ζ forms part of the T cell receptor (TCR where it plays an important role in coupling antigen recognition to several intracellular signal-transduction pathways leading to T cell effector functions. Down regulation of CD3ζ leads to impairment of immune responses including reduced cell proliferation and cytokine production. In experimental models helminth parasites have been shown to modulate immune responses directed against them and unrelated antigens, so called bystander antigens, but there is a lack of studies validating these observations in humans. This study focused on investigated the relationship between expression levels of the TCR CD3ζ chain with lymphocyte cell proliferation during human infection with the helminth parasite, Schistosoma haematobium which causes uro-genital schistosomiasis. Using flow cytometry, peripheral blood mononuclear cells (PBMCs from individuals naturally exposed to S. haematobium in rural Zimbabwe were phenotyped, and expression levels of CD3ζ on T cells were related to intensity of infection. In this population, parasite infection intensity was inversely related to CD3ζ expression levels (p<0.05, consistent with down-regulation of CD3ζ expression during helminth infection. Furthermore, PBMC proliferation was positively related to expression levels of CD3ζ (p<0.05 after allowing for confounding variables (host age, sex, infection level. CD3ζ expression levels had a differing relationship between immune correlates of susceptibility and immunity, measured by antibody responses, indicating a complex relationship between immune activation status and immunity. The relationships between the CD3ζ chain of the TCR and schistosome infection, PBMC proliferation and schistosome-specific antibody responses have not previously been reported, and these results may indicate a mechanism for the impaired T cell proliferative responses observed during human schistosome infection.

  3. Down-regulation of transcobalamin receptor TCblR/CD320 by siRNA inhibits cobalamin uptake and proliferation of cells in culture

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    Lai, Shao-Chiang [School of Graduate Studies, SUNY-Downstate Medical Center, Brooklyn, NY 11203 (United States); Nakayama, Yasumi; Sequeira, Jeffrey M. [Department of Medicine, Cell Biology, SUNY-Downstate Medical Center, Brooklyn, NY 11203 (United States); Quadros, Edward V., E-mail: Edward.Quadros@downstate.edu [Department of Medicine, Cell Biology, SUNY-Downstate Medical Center, Brooklyn, NY 11203 (United States); School of Graduate Studies, SUNY-Downstate Medical Center, Brooklyn, NY 11203 (United States)

    2011-07-01

    The clinical phenotype of cobalamin (Cbl) deficiency is dictated by the essential role of this vitamin in two key enzymatic reactions. Multiple proteins and receptors participate in the absorption, transport and delivery of this vitamin to tissue cells. Cellular uptake of Cbl is mediated by transcobalamin (TC), a plasma protein and a transmembrane receptor (TCblR) with high affinity for TC saturated with Cbl. Knockdown of TCblR with siRNA results in decreased TC-Cbl uptake. The ensuing Cbl deficiency leads to an increase in doubling time and decreased proliferation of these cells. The study confirms the seminal role of this receptor in the cellular uptake of Cbl and its down-regulation as a potential strategy to inhibit proliferation of cancer cells.

  4. IL-13 promotes collagen accumulation in Crohn's disease fibrosis by down-regulation of fibroblast MMP synthesis: a role for innate lymphoid cells?

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    Jennifer R Bailey

    Full Text Available BACKGROUND: Fibrosis is a serious consequence of Crohn's disease (CD, often necessitating surgical resection. We examined the hypothesis that IL-13 may promote collagen accumulation within the CD muscle microenvironment. METHODS: Factors potentially modulating collagen deposition were examined in intestinal tissue samples from fibrotic (f CD and compared with cancer control (C, ulcerative colitis (UC and uninvolved (u CD. Mechanisms attributable to IL-13 were analysed using cell lines derived from uninvolved muscle tissue and tissue explants. RESULTS: In fCD muscle extracts, collagen synthesis was significantly increased compared to other groups, but MMP-2 was not co-ordinately increased. IL-13 transcripts were highest in fCD muscle compared to muscle from other groups. IL-13 receptor (R α1 was expressed by intestinal muscle smooth muscle, nerve and KIR(+ cells. Fibroblasts from intestinal muscle expressed Rα1, phosphorylated STAT6 in response to IL-13, and subsequently down-regulated MMP-2 and TNF-α-induced MMP-1 and MMP-9 synthesis. Cells with the phenotype KIR(+CD45(+CD56(+/-CD3(- were significantly increased in fCD muscle compared to all other groups, expressed Rα1 and membrane IL-13, and transcribed high levels of IL-13. In explanted CD muscle, these cells did not phosphorylate STAT6 in response to exogenous IL-13. CONCLUSIONS: The data indicate that in fibrotic intestinal muscle of Crohn's patients, the IL-13 pathway is stimulated, involving a novel population of infiltrating IL-13Rα1(+, KIR(+ innate lymphoid cells, producing IL-13 which inhibits fibroblast MMP synthesis. Consequently, matrix degradation is down-regulated and this leads to excessive collagen deposition.

  5. Budesonide suppresses pulmonary antibacterial host defense by down-regulating cathelicidin-related antimicrobial peptide in allergic inflammation mice and in lung epithelial cells

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    Wang Peng

    2013-02-01

    Full Text Available Abstract Background Glucocorticoids are widely regarded as the most effective treatment for asthma. However, the direct impact of glucocorticoids on the innate immune system and antibacterial host defense during asthma remain unclear. Understanding the mechanisms underlying this process is critical to the clinical application of glucocorticoids for asthma therapy. After sensitization and challenge with ovalbumin (OVA, BALB/c mice were treated with inhaled budesonide and infected with Pseudomonas aeruginosa (P. aeruginosa. The number of viable bacteria in enflamed lungs was evaluated, and levels of interleukin-4 (IL-4 and interferon-γ (IFN-γ in serum were measured. A lung epithelial cell line was pretreated with budesonide. Levels of cathelicidin-related antimicrobial peptide (CRAMP were measured by immunohistochemistry and western blot analysis. Intracellular bacteria were observed in lung epithelial cells. Results Inhaled budesonide enhanced lung infection in allergic mice exposed to P. aeruginosa and increased the number of viable bacteria in lung tissue. Higher levels of IL-4 and lower levels of IFN-γ were observed in the serum. Budesonide decreased the expression of CRAMP, increased the number of internalized P. aeruginosa in OVA-challenged mice and in lung epithelial cell lines. These data indicate that inhaled budesonide can suppress pulmonary antibacterial host defense by down-regulating CRAMP in allergic inflammation mice and in cells in vitro. Conclusions Inhaled budesonide suppressed pulmonary antibacterial host defense in an asthmatic mouse model and in lung epithelium cells in vitro. This effect was dependent on the down-regulation of CRAMP.

  6. MiR-29b inhibits collagen maturation in hepatic stellate cells through down-regulating the expression of HSP47 and lysyl oxidase

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    Zhang, Yifei; Ghazwani, Mohammed; Li, Jiang [Center for Pharmacogenetics, Department of Pharmaceutical Sciences, School of Pharmacy, University of Pittsburgh, Pittsburgh, PA 15261 (United States); Sun, Ming; Stolz, Donna B. [Department of Cell Biology and Physiology, School of Medicine, University of Pittsburgh, Pittsburgh, PA 15261 (United States); He, Fengtian [Department of Biochemistry and Molecular Biology, College of Basic Medical Sciences, Third Military Medical University, Chongqing 400038 (China); Fan, Jie [Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15261 (United States); Xie, Wen [Center for Pharmacogenetics, Department of Pharmaceutical Sciences, School of Pharmacy, University of Pittsburgh, Pittsburgh, PA 15261 (United States); Li, Song, E-mail: sol4@pitt.edu [Center for Pharmacogenetics, Department of Pharmaceutical Sciences, School of Pharmacy, University of Pittsburgh, Pittsburgh, PA 15261 (United States)

    2014-04-18

    Highlights: • Enhanced HSP47 and LOX expression is associated with decreased miR-29b level in liver fibrosis. • miR-29b down-regulates HSP47 and LOX expression. • The suppression of HSP47 and LOX by miR-29b is mediated by putative sites at their 3′-UTRs. • miR-29b inhibits extracellular LOX activity and collagen maturation. - Abstract: Altered expression of miR-29b is implicated in the pathogenesis and progression of liver fibrosis. We and others previously demonstrated that miR-29b down-regulates the expression of several extracellular-matrix (ECM) genes including Col 1A1, Col 3A1 and Elastin via directly targeting their 3′-UTRs. However, whether or not miR-29b plays a role in the post-translational regulation of ECM biosynthesis has not been reported. Heat shock protein 47 (HSP47) and lysyl oxidase (LOX) are known to be essential for ECM maturation. In this study we have demonstrated that expression of HSP47 and LOX was significantly up-regulated in culture-activated primary rat hepatic stellate cells (HSCs), TGF-β stimulated LX-2 cells and liver tissue of CCl{sub 4}-treated mice, which was accompanied by a decrease of miR-29b level. In addition, over-expression of miR-29b in LX-2 cells resulted in significant inhibition on HSP47 and LOX expression. Mechanistically, miR-29b inhibited the expression of a reporter gene that contains the respective full-length 3′-UTR from HSP47 and LOX gene, and this inhibitory effect was abolished by the deletion of a putative miR-29b targeting sequence from the 3′-UTRs. Transfection of LX-2 cells with miR-29b led to abnormal collagen structure as shown by electron-microscopy, presumably through down-regulation of the expression of molecules involved in ECM maturation including HSP47 and LOX. These results demonstrated that miR-29b is involved in regulating the post-translational processing of ECM and fibril formation.

  7. Enterococcus faecalis Infection and Reactive Oxygen Species Down-Regulates the miR-17-92 Cluster in Gastric Adenocarcinoma Cell Culture

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    Jesper A. B. Strickertsson

    2014-08-01

    Full Text Available Chronic inflammation due to bacterial overgrowth of the stomach predisposes to the development of gastric cancer and is also associated with high levels of reactive oxygen species (ROS. In recent years increasing attention has been drawn to microRNAs (miRNAs due to their role in the pathogenesis of many human diseases including gastric cancer. Here we studied the impact of infection by the gram-positive bacteria Enterococcus faecalis (E. faecalis on global miRNA expression as well as the effect of ROS on selected miRNAs. Human gastric adenocarcinoma cell line MKN74 was infected with living E. faecalis for 24 h or for 5 days or with E. faecalis lysate for 5 days. The miRNA expression was examined by microarray analysis using Affymetrix GeneChip miRNA Arrays. To test the effect of ROS, MKN74 cells were treated with 100 mM tert-Butyl hydroperoxide (TBHP. Following 5 days of E. faecalis infection we found 91 differentially expressed miRNAs in response to living bacteria and 2 miRNAs responded to E. faecalis lysate. We verified the down-regulation of the miR-17-92 and miR-106-363 clusters and of other miRNAs involved in the oxidative stress-response by qRT-PCR. We conclude that only infection by living E. faecalis bacteria caused a significant global response in miRNA expression in the MKN74 cell culture. E. faecalis infection as well as ROS stimulation down-regulated the expression of the miR-17-92 cluster. We believe that these changes could reflect a general response of gastric epithelial cells to bacterial infections.

  8. P120ctn overexpression enhances β-catenin-E-cadherin binding and down regulates expression of survivin and cyclin D1 in BEL-7404 hepatoma cells

    Institute of Scientific and Technical Information of China (English)

    Chao-Zan Nong; Li-Li Pan; Wei-Sheng He; Xi-Liang Zha; Hai-Hong Ye; Hua-Yi Huang

    2006-01-01

    AIM: To understand the role of P120ctn in E-cadherin-mediated cell-cell adhesion and signaling as well as in hepatoma cell biological function.METHODS: We stably overexpressed p120ctn isoform 3A in BEL-7404 human hepatoma cells and studied the effect of p120ctn on β-catenin and E-cadherin binding as well as p120ctn and β-catenin subcellular localization using immunoprecipitation, Western blotting and confocalmicroscopy. We also investigated the inhibitory effect of p120ctn transfection on the expression of apoptotic protein survivin survivin and cell cycle regulator cyclin D1in the cells.RERULTS: Western blotting indicated that p120ctn expression increased after cells were transfected with p120ctn isoform 3A. The protein was located mainly at membrane under immunofluorescent microscope.β-catenin nuclear expression was reduced after overexpression of p120ctn isoform 3A. The p120ctn-E-cadherin binding increased after transfection of p120ctn isoform 3A. Furthermore, overexpression of p120ctn down regulated the expression of apoptotic protein survivin and cell cycle regulator cyclin D1. These effects led to reduction of cell proliferation.CONCLUSION: Our results indicate that p120ctn plays an important role in regulating the formation of E-cadherin and -catenin complex, cell apoptosis, cell cycle and cancer cell biological function.

  9. The molecular pathway of low concentration of sodium arsenite in inducing differentiation of liver cancer stem cells by down-regulating promyelocytic leukemia protein expression

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    Shi-long JIN

    2016-01-01

    Full Text Available Objective  To study the molecular pathway of low concentration of sodium arsenite in inducing differentiation of liver cancer stem cells. Methods  Western blotting analysis, immunofluorescence assay and quantitative PCR were used to examine the gene and protein expression of promyelocytic leukemia (PML, Oct4 and Sox2 in HCC tissue and cell lines, and the molecule pathway of low concentration of sodium arsenite inducing differentiation of liver cancer stem cells was confirmed by comparing the changes in the gene and protein expression of PML,Oct4 and Sox2 in HCC cells and biological function of LCSCs after the treatment with low concentration of sodium arsenite. Results  0.5μg/ml of sodium arsenite was shown to alter the biological characteristics of LCSCs in HuH7 and primary HCC cells, including the ability to form tumor spheres, resistance to pirarubicin (P<0.01, and the capability of forming tumors after allogeneic transplantation (P<0.05. Both HCC cells and tissues expressed the gene and protein of PML,Oct4 and Sox2, and 0.5μg/ml of sodium arsenite not only downregulated the gene and protein expression of Oct4 (P<0.05 and Sox2 in HCC cells (P<0.05, but also downregulated the protein expression of PML (P<0.05. In contrast, sodium arsenite did not inhibit the gene expression of PML in Hep3B, HepG2, SMCC-7721, HuH7 and primary HCC cells. Furthermore, through down-regulated PML protein expression with arsenite, the biological characteristics of HuH7 and primary HCC cells containing LCSCs was simultaneously altered, and the expression of stem gene Oct4 and Sox2 was downregulated (P<0.05, while HCC cells proliferation was inhibited as well. Conclusions  Both HCC tissues and cells can express the PML gene and PML protein. Low concentrations of sodium arsenite would directly bind to PML protein in HCC cells, resulting in degradation of the PML protein, followed by collapse of PML-NBs, inhibition of transcription of the proliferation

  10. Down-regulation of B cell-related genes in peripheral blood leukocytes of Parkinson's disease patients with and without GBA mutations.

    Science.gov (United States)

    Kobo, Hila; Bar-Shira, Anat; Dahary, Dvir; Gan-Or, Ziv; Mirelman, Anat; Goldstein, Orly; Giladi, Nir; Orr-Urtreger, Avi

    2016-02-01

    Parkinson's disease (PD) is a common neurodegenerative disorder, caused by aging, genetic and environmental factors. Many genes and genetic loci have been implicated in autosomal dominant and recessive PD, among them SNCA, LRRK2, GBA, Parkin, DJ1 and PINK1. Mutations in the LRRK2 and GBA genes are especially common among PD patients of Ashkenazi-Jewish (AJ) origin, accounting for over a third of the patient population. We aimed to identify genes and cellular pathways that may be involved in GBA-associated PD. Whole genome expression analysis was performed using peripheral blood leukocytes (PBLs) of PD patients with mutations in the GBA gene (PD-GBA, n = 59) compared to healthy controls (n = 59). Significant expression changes were detected in 26 genes, most of them were down-regulated in patients and annotated to B cell or immune-related functions. The expression levels of five membrane-bound B cell genes (FCRL1, CD19, CD22, CD79A and CD180) were further analyzed in four distinct populations: (1) Healthy controls (n = 20), (2) PD-GBA (n = 20), (3) PD patients who do not carry LRRK2 or GBA mutations (PD-NC, n = 20), (4) Asymptomatic 1st degree family members, with (n = 15) or without (n = 15) GBA mutations. In qRT-PCR analysis, all five genes were down-regulated in patients (PD-GBA and PD-NC) compared to controls. These changes in expression were not observed when comparing family members who carry GBA mutations to non-carrier family members. Furthermore, these expression levels were disease-duration dependent: the most significant decreased expression occurred after the first two years of onset, and remained steady after 6 years. These results further support the involvement of B cell-related genes in PD and correlate the level of reduced expression to disease duration.

  11. Andrographolide down-regulates hypoxia-inducible factor-1α in human non-small cell lung cancer A549 cells.

    Science.gov (United States)

    Lin, Hui-Hsuan; Tsai, Chia-Wen; Chou, Fen-Pi; Wang, Chau-Jong; Hsuan, Shu-Wen; Wang, Cheng-Kun; Chen, Jing-Hsien

    2011-02-01

    Andrographolide (Andro), a diterpenoid lactone isolated from a traditional herbal medicine Andrographis paniculata, is known to possess multiple pharmacological activities. In our previous study, Andro had been shown to inhibit non-small cell lung cancer (NSCLC) A549 cell migration and invasion via down-regulation of phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway. Here we demonstrated that Andro inhibited the expression of hypoxia-inducible factor-1α (HIF-1α) in A549 cells. HIF-1α plays an important role in tumor growth, angiogenesis and lymph node metastasis of NSCLC. The Andro-induced decrease of cellular protein level of HIF-1α was correlated with a rapid ubiquitin-dependent degradation of HIF-1α, and was accompanied by increased expressions of hydroxyl-HIF-1α and prolyl hydroxylase (PHD2), and a later decrease of vascular endothelial growth factor (VEGF) upon the treatment of Andro. The Andro-inhibited VEGF expression appeared to be a consequence of HIF-1α inactivation, because its DNA binding activity was suppressed by Andro. Molecular data showed that all these effects of Andro might be mediated via TGFβ1/PHD2/HIF-1α pathway, as demonstrated by the transfection of TGFβ1 overexpression vector and PHD2 siRNA, and the usage of a pharmacological MG132 inhibitor. Furthermore, we elucidated the involvement of Andro in HIF-1α transduced VEGF expression in A549 cells and other NSCLC cell lines. In conclusion, these results highlighted the potential effects of Andro, which may be developed as a chemotherapeutic or an anti-angiogenesis agent for NSCLC in the future.

  12. Matrine promotes G0/G1 arrest and down-regulates cyclin D1 expression in human rhabdomyosarcoma cells.

    Science.gov (United States)

    Guo, L; Xue, T Y; Xu, W; Gao, J Z

    2013-09-01

    Matrine has a broad-spectrum of anti-cancer effects and is efficient in the inhibition of proliferation of hepatoma cells, leukemia cells and neuroblastoma cell. However, its efficacy and tentative mechanisms in rhabdomyosarcoma have not been addressed before. This study aimed to investigate the effects of Matrine on cell cycle and expression of cyclin D1 in human rhabdomyosarcoma cells (RD cell line). RD cell line was treated with different concentrations (0, 0.5, 1.0, and 1.5 mg/mL) of Matrine, and cell proliferation and cell cycle were evaluated using, respectively, MTT assay and flow cytometry. The effect of Matrine on cyclin D1 mRNA levels was measured by RT-PCR. There was a dose-dependent inhibition of proliferation in the matrine-treated group (inhibition of proliferation rate in control cells 12.70 ± 0.35%; Matrine-treated cells [0.5, 1.0, and 1.5 mg/mL]: 31.16 ± 0.11%, 42.96 ± 0.9%, and 57.26 ± 0.8%). The G0 / G1 ratio in study groups were, respectively, 58.44 ± 3.57%, 64.79 ± 2.03%, 69.97 ± 2.89% and 75.03 ± 1.23%.Cyclin D1 mRNA levels progressively diminished (control group ratio of cyclin D1 / β-actin: 0.59 ± 0.06; Matrine: 0.35 ± 0.05, 0.27 ± 0.02 and 0.04 ± 0.03). All aforementioned changes were significant (PMatrine markedly suppresses cell proliferation in RD cells by decreasing expression of cyclin D1 mRNA and blocking the cell cycle at the G0 / G1 stage.

  13. Low-amplitude high frequency vibration down-regulates myostatin and atrogin-1 expression, two components of the atrophy pathway in muscle cells.

    Science.gov (United States)

    Ceccarelli, Gabriele; Benedetti, Laura; Galli, Daniela; Prè, Deborah; Silvani, Giulia; Crosetto, Nicola; Magenes, Giovanni; Cusella De Angelis, Maria Gabriella

    2014-05-01

    Whole body vibration (WBV) is a very widespread mechanical stimulus used in physical therapy, rehabilitation and fitness centres. It has been demonstrated that vibration induces improvements in muscular strength and performance and increases bone density. We investigated the effects of low-amplitude, high frequency vibration (HFV) at the cellular and tissue levels in muscle. We developed a system to produce vibrations adapted to test several parameters in vitro and in vivo. For in vivo experiments, we used newborn CD1 wild-type mice, for in vitro experiments, we isolated satellite cells from 6-day-old CD1 mice, while for proliferation studies, we used murine cell lines. Animals and cells were treated with high frequency vibration at 30 Hz. We analyzed the effects of mechanical stimulation on muscle hypertrophy/atrophy pathways, fusion enhancement of myoblast cells and modifications in the proliferation rate of cells. Results demonstrated that mechanical vibration strongly down-regulates atrophy genes both in vivo and in vitro. The in vitro experiments indicated that mechanical stimulation promotes fusion of satellite cells treated directly in culture compared to controls. Finally, proliferation experiments indicated that stimulated cells had a decreased growth rate compared to controls. We concluded that vibration treatment at 30 Hz is effective in suppressing the atrophy pathway both in vivo and in vitro and enhances fusion of satellite muscle cells.

  14. Down-regulation of hypoxia-inducible factor-1 suppresses malignant biological behavior of triple-negative breast cancer cells.

    Science.gov (United States)

    Wang, Fang; Chang, Miaomiao; Shi, Yonghong; Jiang, Lili; Zhao, Jing; Hai, Ling; Sharen, Gaowa; Du, Hua

    2014-01-01

    This study is to investigate the effect and mechanism of reduced hypoxia-inducible factor (HIF)-1a expression on malignant behavior of MDA-MB-231 cells. HIF-1α expression was interfered by siRNA. Western blot was used to detect protein expression of HIF-1α, active fragments of caspase 3 and vimentin. Cell count, flow cytometry and Hoechst staining were used to evaluate cell growth and apoptosis. Matrigel invasion and wound scratch assay were performed to measure the ability of cell invasion and migration. After MDA-MB-231 cells were transfected with HIF-1α-targeted siRNA, HIF-1α protein expression was successfully interrupted and cell growth was retarded. Compared with random siRNA group, reduced HIF-1α protein expression in HIF-1α-targeted siRNA group facilitated cell apoptosis but had no effect on cell cycle. In addition, cells treated with HIF-1α-targeted siRNA expressed active fragments of caspase 3 (17 and 12 kD) after serum starvation for 0 to 60 h. Caspase 3 activity assay further confirmed the above finding. Reduced HIF-1α expression impaired the migration and invasiveness with a reduction in the expression of vimentin and CK18 protein. Inhibition of HIF-1α protein synthesis or enhancement of its degradation reversed its malignant phenotypes and could probably be a potential means for the treatment of triple-negative breast cancer.

  15. Down-regulation of DNA topoisomerase IIalpha leads to prolonged cell cycle transit in G2 and early M phases and increased survival to microtubule-interacting agents.

    Science.gov (United States)

    Skladanowski, Andrzej; Côme, Marie-George; Sabisz, Michal; Escargueil, Alexandre E; Larsen, Annette K

    2005-09-01

    Microtubule binders are cell cycle-specific agents with preferential cytotoxicity toward mitotic cells. We have characterized vincristine-selected human leukemia cells to establish whether development of vincristine resistance was accompanied by changes in cell cycle kinetics and distribution. Our results indicate that vincristine resistance is accompanied by delayed G2 transit and prolonged early mitosis in both the absence and the presence of the microtubule binder nocodazole. The altered G2/M regulation is accompanied by resistance to short-term (12 h) but not continuous nocodazole exposure in agreement with the transient nature of the observed cell cycle alterations. Western blot analysis indicates that vincristine-selection is accompanied by down-regulation of topoisomerase IIalpha without detectable alterations of the other mitotic regulators studied, including Cdk1, p21, 14-3-3sigma, and 14-3-3epsilon. This was associated with at least 7-fold less chromosome-associated topoisomerase IIalpha, decreased catalytic activity, and cross-resistance to topoisomerase II inhibitors. Characterization of isogenic cell lines expressing different levels of topoisomerase II proteins shows that cellular levels of topoisomerase IIalpha, but not the closely related topoisomerase IIbeta, directly influence the cell cycle kinetics in G2 and early mitosis as well as the resistance to nocodazole. These results underline the importance of topoisomerase IIalpha in late G2 and early M phases and provide evidence for an as-yet-unsuspected interaction between topoisomerase II and microtubule-directed agents.

  16. Down-regulation of PKHD1 induces cell apoptosis through PI3K and NF-{kappa}B pathways

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    Sun, Liping [Division of Nephrology, Shenzhen People' s Hospital, Second Clinical Medical College, Jinan University, Shenzhen 518020 (China); Wang, Shixuan [Renal Division, Brigham and Women' s Hospital and Harvard Medical School, Boston, MA 02115 (United States); Hu, Chaofeng [Department of Pathophysiology, Medical College, Jinan University, Guangzhou, 510632 (China); Zhang, Xinzhou, E-mail: slp08@126.com [Division of Nephrology, Shenzhen People' s Hospital, Second Clinical Medical College, Jinan University, Shenzhen 518020 (China)

    2011-04-15

    Mutations in PKHD1 (polycystic kidney and hepatic disease gene 1) gene cause the autosomal recessive polycystic kidney disease (ARPKD). Fibrocystin/polyductin (FPC), encoded by PKHD1, is a membrane-associated receptor-like protein. Although it is widely accepted that cystogenesis is mostly due to aberrant cell proliferation and apoptosis, it is still unclear how apoptosis is regulated. The aim of this study is to analyze the relationship among apoptosis, phosphatidylinositol 3-kinase (PI3K)/Akt and nuclear factor {kappa}B (NF-{kappa}B) in FPC knockdown kidney cells. We show that PKHD1-silenced HEK293 cells demonstrate a higher PI3K/Akt activity. Selective inhibition of PI3K/Akt using LY294002 or wortmannin in these cells increases serum starvation-induced HEK293 cell apoptosis with a concomitant decrease in cell proliferation and higher caspase-3 activity. PI3K/Akt inhibition also leads to increased NF-{kappa}B activity in these cells. We conclude that the PI3K/Akt pathway is involved in apoptotic function in PKHD1-silenced cells, and PI3K/Akt inhibition correlates with upregulation of NF-{kappa}B activity. These observations provide a potential platform for determining FPC function and therapeutic investigation of ARPKD.

  17. p38 MAPK-Mediated Bmi-1 down-regulation and defective proliferation in ATM-deficient neural stem cells can be restored by Akt activation.

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    Jeesun Kim

    Full Text Available A-T (ataxia telangiectasia is a genetic disease caused by a mutation in the Atm (A-T mutated gene that leads to neurodegeneration. Despite an increase in the numbers of studies in this area in recent years, the mechanisms underlying neurodegeneration in human A-T are still poorly understood. Previous studies demonstrated that neural stem cells (NSCs isolated from the subventricular zone (SVZ of Atm(-/- mouse brains show defective self-renewal and proliferation, which is accompanied by activation of chronic p38 mitogen-activated protein kinase (MAPK and a lower level of the polycomb protein Bmi-1. However, the mechanism underlying Bmi-1 down-regulation and its relevance to defective proliferation in Atm(-/- NSCs remained unclear. Here, we show that over-expression of Bmi-1 increases self-renewal and proliferation of Atm(-/- NSCs to normal, indicating that defective proliferation in Atm(-/- NSCs is a consequence of down-regulation of Bmi-1. We also demonstrate that epidermal growth factor (EGF-induced Akt phosphorylation renders Bmi-1 resistant to the proteasomal degradation, leading to its stabilization and accumulation in the nucleus. However, inhibition of the Akt-dependent Bmi-1 stabilizing process by p38 MAPK signaling reduces the levels of Bmi-1. Treatment of the Atm(-/- NSCs with a specific p38 MAPK inhibitor SB203580 extended Bmi-1 posttranscriptional turnover and H2A ubiquitination in Atm(-/- NSCs. Our observations demonstrate the molecular basis underlying the impairment of self-renewal and proliferation in Atm(-/- NSCs through the p38 MAPK-Akt-Bmi-1-p21 signaling pathway.

  18. Down-regulation of microRNA-181b is a potential prognostic marker of non-small cell lung cancer.

    Science.gov (United States)

    Yang, Junquan; Liu, Hongxia; Wang, Hongbin; Sun, Yuman

    2013-08-01

    The aim of this study was to investigate the clinical significance of microRNA-181b (miR-181b) expression in non-small cell lung cancer (NSCLC). MiR-181b expression in 126 pairs of surgically removed NSCLC tissues and their corresponding normal lung tissues was measured by real-time quantitative RT-PCR assay. Additionally, the correlation of miR-181b expression with clinicopathological factors or prognosis of patients was analyzed. At first, miR-181b expression was significantly down-regulated in NSCLC tissues as compared with their normal counterparts (PmiR-181b expression was found to be closely correlated with larger tumor size (P=0.02), higher p-TNM stage (P=0.008) and positive lymph node metastasis (P=0.03) of NSCLC patients. After that, survival analysis found that the overall survival (P=0.001) and disease-free survival (P=0.008) of NSCLC patients with low miR-181b expression were both significantly poorer compared to those patients with high miR-181b expression. Finally, both univariate and multivariate analyses demonstrated that low miR-181b expression may be a poor prognostic marker of NSCLC patients. This is the first study to indicate that down-regulation of miR-181b may be correlated with aggressive disease progression and poor prognosis of NSCLC patients, suggesting that miR-181b might be involved in lung carcinogenesis and become a potential prognostic marker for NSCLC.

  19. Growth inhibition of human breast cancer cells and down-regulation of ODC1 and ADA genes by Nepeta binaloudensis

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    Akbar Safipour Afshar

    Full Text Available ABSTRACT Nepeta binaloudensis Jamzad, Lamiaceae, is a rare medicinal plant endemic to Iran. In spite of many studies about the chemical constituents and antibacterial effects of this species, no report has been provided about its cytotoxic and anticancer activities. In this study we have evaluated the effects of EtOH 70%, hexane and aqueous extracts of N. binaloudensis on the cell proliferation and n-hexane extract on the expression of adenosine deaminase and ornithine decarboxylase 1 genes in breast cancer cell lines (MCF-7, MDA-MB-231 compared to non-cancer line (MCF-10A. The cell lines were subjected to increasing doses of the extracts ranging from 10 to 320 µg/ml. Cell viability was quantified by MTS assay. Expression of adenosine deaminase and ornithine decarboxylase 1 genes was analyzed by real time PCR. N. binaloudensis inhibited the growth of malignant cells in a time and dose-dependent manner. Among extracts of N. binaloudensis, the hexane extract was found to be more toxic compared to other extracts. Results showed a marked decrease in the expression of ornithine decarboxylase 1 and adenosine deaminase genes in cancer cell lines. At 60 µg/ml concentration of N. binaloudensis hexane extract ornithine decarboxylase 1 and adenosine deaminase mRNA expression were reduced 4.9 fold and 3.5 fold in MCF-7 cell line and 3.6 fold and 2.6 fold in MDA-MB-231 cell line compared to control, respectively. The result of our study highlights the potential influences of N. binaloudensis hexane extract on ornithine decarboxylase 1 and adenosine deaminase genes expression in breast cancer cells and its relation to inhibition of cancer cell growth.

  20. Down-Regulation of miR-92 in Breast Epithelial Cells and in Normal but Not Tumour Fibroblasts Contributes to Breast Carcinogenesis.

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    Laura Smith

    Full Text Available MicroRNA (miR expression is commonly dysregulated in many cancers, including breast. MiR-92 is one of six miRs encoded by the miR-17-92 cluster, one of the best-characterised oncogenic miR clusters. We examined expression of miR-92 in the breast epithelium and stroma during breast cancer progression. We also investigated the role of miR-92 in fibroblasts in vitro and showed that down-regulation in normal fibroblasts enhances the invasion of breast cancer epithelial cells.We used laser microdissection (LMD to isolate epithelial cells from matched normal, DCIS and invasive tissue from 9 breast cancer patients and analysed miR-92 expression by qRT-PCR. Expression of ERβ1, a direct miR-92 target, was concurrently analysed for each case by immunohistochemistry. LMD was also used to isolate matched normal (NFs and cancer-associated fibroblasts (CAFs from 14 further cases. Effects of miR-92 inhibition in fibroblasts on epithelial cell invasion in vitro was examined using a Matrigel™ assay. miR-92 levels decreased in microdissected epithelial cells during breast cancer progression with highest levels in normal breast epithelium, decreasing in DCIS (p<0.01 and being lowest in invasive breast tissue (p<0.01. This was accompanied by a shift in cell localisation of ERβ1 from nuclear expression in normal breast epithelium to increased cytoplasmic expression during progression to DCIS (p = 0.0078 and invasive breast cancer (p = 0.031. ERβ1 immunoreactivity was also seen in stromal fibroblasts in tissues. Where miR-92 expression was low in microdissected NFs this increased in matched CAFs; a trend also seen in cultured primary fibroblasts. Down-regulation of miR-92 levels in NFs but not CAFs enhanced invasion of both MCF-7 and MDA-MB-231 breast cancer epithelial cells.miR-92 is gradually lost in breast epithelial cells during cancer progression correlating with a shift in ERβ1 immunoreactivity from nuclei to the cytoplasm. Our data support a functional

  1. Down-regulation of mTOR leads to up-regulation of osteoprotegerin in bone marrow cells

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    Mogi, Makio, E-mail: makio@dpc.aichi-gakuin.ac.jp [Department of Medicinal Biochemistry, School of Pharmacy, Aichi-Gakuin University, 1-100 Kusumoto, Chikusa, Nagoya, Aichi 464-8650 (Japan); Kondo, Ayami [Department of Medicinal Biochemistry, School of Pharmacy, Aichi-Gakuin University, 1-100 Kusumoto, Chikusa, Nagoya, Aichi 464-8650 (Japan)

    2009-06-19

    Osteoprotegerin (OPG)/osteoclastogenesis inhibitory factor regulates bone mass by inhibiting osteoclastic bone resorption. mTOR, which is the mammalian target of rapamycin, is a kinase and central regulator of cell growth, proliferation, and survival. By using Rapamycin, we studied whether mTOR pathway is associated with OPG protein production in the mouse bone marrow-derived stromal cell line ST2. Rapamycin markedly increased the level of soluble OPG in ST2 cells. This antibiotic treatment resulted in the suppression of phosphorylation of mTOR. Rapamycin had no effects on the proliferation, differentiation, or apoptosis of the cells. Treatment with bone morphogenetic protein-4, which can induce OPG protein in ST2 cells, also resulted in a decrease in the density of the phospho-mTOR-band, suggesting that the suppression of the phospho-mTOR pathway is necessary for OPG production in ST2 cells. Thus, suitable suppression of mTOR phosphorylation is a necessary requirement for OPG production in bone marrow stromal cells.

  2. Overexpression of HTRA1 leads to down-regulation of fibronectin and functional changes in RF/6A cells and HUVECs.

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    Jingjing Jiang

    Full Text Available Multiple genetic studies have suggested that high-temperature requirement serine protease (HTRA1 is associated with polypoidal choroidal vasculopathy (PCV. To date, no functional studies have investigated the biological effect of HTRA1 on vascular endothelial cells, essential vascular components involved in polypoidal vascular abnormalities and arteriosclerosis-like changes. In vitro studies were performed to investigate the effect of HTRA1 on the regulation of fibronectin, laminin, vascular endothelial growth factor (VEGF, platelet derived growth factor receptor (PDGFR and matrix metalloparoteinases 2 (MMP-2 and the role of HTRA1 in choroid-retina endothelial (RF/6A and human umbilical vein endothelial (HUVEC cells. Lentivirus-mediated overexpression of HTRA1 was used to explore effects of the protease on RF/6A and HUVEC cells in vitro. HTRA1 overexpression inhibited the proliferation, cell cycle, migration and tube formation of RF/6A and HUVEC cells, effects that might contribute to the early stage of PCV pathological lesions. Fibronectin mRNA and protein levels were significantly down-regulated following the upregulation of HTRA1, whereas the expressions of laminin, VEGF and MMP-2 were unaffected by alterations in HTRA1 expression. The decreased biological function of vascular endothelial cells and the degradation of extracellular matrix proteins, such as fibronectin, may be involved in a contributory role for HTRA1 in PCV pathogenesis.

  3. Cocaine enhances HIV-1 replication in CD4+ T cells by down-regulating MiR-125b.

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    Chinmay K Mantri

    Full Text Available The main objective of this study was to examine effects of cocaine on HIV-1 replication in primary CD4+ T cells. Cocaine a commonly used drug among HIV-1 positive individuals serves as a cofactor for HIV-1 infection and progression to acquired immunodeficiency syndrome (AIDS. Accumulating evidence suggest that cocaine increases HIV-1 replication in cell cultures, peripheral blood mononuclear cells (PBMCs and animal models. Intriguingly, there are no studies on cocaine-induced alterations in HIV-1 replication in primary CD4+ T cells that serve as the main targets for HIV-1 replication in vivo. In this report, we demonstrate cocaine-induced enhancement of HIV-1 replication in primary CD4+ T cells isolated from human PBMCs. To decipher a potential mechanism, we examined whether cocaine targets the innate antiviral immunity of CD4+ T cells mediated by cellular microRNAs (miRNAs. This is because recently a network of anti-HIV miRNAs in CD4+ T cells is highlighted to suppress viral replication. Our genome wide miRNA expression analysis indicated downregulation of several anti-HIV miRNAs (miR-28, miR-125b, miR-150, miR-223, and miR-382 in cocaine treated CD4+ T cells. However, our real-time quantitative PCR analysis revealed significant downregulation of miR-125b only. Our results illustrated that miR-125b knockdown enhances HIV-1 replication, whereas overexpression of miR-125b decreases HIV-1 replication in these cells. Therefore, we believe miR-125b is a key player for the cocaine induced enhancement of HIV-1 replication in CD4+ T cells. Since, miR-125b targets the 3' UTR regions of HIV-1 transcripts and inhibits viral protein translation, our data suggest modulation of post entry steps of HIV-1 by cocaine. Given that a plethora of studies suggest that cocaine regulates HIV entry, our results implicate a potentially novel mechanism by which cocaine can increase viral replication in CD4+ T cells.

  4. Inhibition of Corydalis decumbens Alkaloids on Hydrogen Peroxideinduced Apoptosis of PC12 Cells through Down-regulating Caspase-3 Expression

    Institute of Scientific and Technical Information of China (English)

    YAN Ren-jie; YANG Yi-fang; LUO Yong-ming; WU Chun-zhen

    2011-01-01

    Objective To extract alkaloids from Corydalis decumbens (AsCD) by supercritical CO2 fluid extraction (SFE) and to evaluate protective effects of AsCD against hydrogen peroxide (H2O2)-induced apoptosis in rat PC12 cells.Methods AsCD were extracted by SFE and oxidative damage PC12 cells model was induced by H2O2.The survival rate of the cells was determined by MTT assay; Lactate dehydrogenase release was determined by ultraviolet spectrophotometry; Flow cytometry was used to detect apoptosis; Caspase-3 mRNA and protein were determined by real-time PCR and Western blotting assay,respectively.Results AsCD remarkably reduced the cytotoxicity,prevented membrane damage,and inhibited cell apoptosis.AsCD inhibited Caspase-3 mRNA and protein expression induced by H2O2 in PC12 cells.Conclusion AsCD possess protective effects against H2O2-induced apoptosis in PC12 cells,and the mechanism of AsCD responsible to the inhibition of apoptosis is possibly attributed to thedown-regulating Caspase-3 expression.AsCD might be useful in the treatment of oxidative stress-related neurodegenerative diseases.

  5. Down Regulation of CIAPIN1 Reverses Multidrug Resistance in Human Breast Cancer Cells by Inhibiting MDR1

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    Xuemei Wang

    2012-06-01

    Full Text Available Cytokine-induced apoptosis inhibitor 1 (CIAPIN1, initially named anamorsin, a newly indentified antiapoptotic molecule is a downstream effector of the receptor tyrosine kinase-Ras signaling pathway. Current study has revealed that CIAPIN1 may have wide and important functions, especially due to its close correlations with malignant tumors. However whether or not it is involved in the multi-drug resistance (MDR process of breast cancer has not been elucidated. To explore the effect of CIAPIN1 on MDR, we examined the expression of P-gp and CIAPIN1 by immunohistochemistry and found there was positive correlation between them. Then we successfully interfered with RNA translation by the infection of siRNA of CIAPIN1 into MCF7/ADM breast cancer cell lines through a lentivirus, and the expression of the target gene was significantly inhibited. After RNAi the drug resistance was reduced significantly and the expression of MDR1mRNA and P-gp in MCF7/ADM cell lines showed a significant decrease. Also the expression of P53 protein increased in a statistically significant way (p ≤ 0.01 after RNAi exposure. In addition, flow cytometry analysis reveals that cell cycle and anti-apoptotic enhancing capability of cells changed after RNAi treatment. These results suggested CIAPIN1 may participate in breast cancer MDR by regulating MDR1 and P53 expression, changing cell cycle and enhancing the anti-apoptotic capability of cells.

  6. Ganodermanontriol (GDNT) exerts its effect on growth and invasiveness of breast cancer cells through the down-regulation of CDC20 and uPA

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    Jiang, Jiahua; Jedinak, Andrej [Cancer Research Laboratory, Methodist Research Institute, Indiana University Health, Indianapolis, IN (United States); Sliva, Daniel, E-mail: dsliva@iuhealth.org [Cancer Research Laboratory, Methodist Research Institute, Indiana University Health, Indianapolis, IN (United States); Department of Medicine, School of Medicine, Indiana University, Indianapolis, IN (United States); Indiana University Simon Cancer Center, School of Medicine, Indiana University, Indianapolis, IN (United States)

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer Ganodermanontriol (GDNT), a Ganoderma mushroom alcohol, inhibits growth of breast cancer cells. Black-Right-Pointing-Pointer CDC20 is over-expressed in tumors but not in the tumor surrounding tissue in breast cancer patients. Black-Right-Pointing-Pointer GDNT inhibits expression of CDC20 in breast cancer cells. Black-Right-Pointing-Pointer GDNT inhibits cell adhesion, cell migration and cell invasion of breast cancer cells. Black-Right-Pointing-Pointer GDNT inhibits secretion of uPA and down-regulates expression of uPAR in breast cancer cells. -- Abstract: Ganoderma lucidum is a medicinal mushroom that has been recognized by Traditional Chinese Medicine (TCM). Although some of the direct anticancer activities are attributed to the presence of triterpenes-ganoderic and lucidenic acids-the activity of other compounds remains elusive. Here we show that ganodermanontriol (GDNT), a Ganoderma alcohol, specifically suppressed proliferation (anchorage-dependent growth) and colony formation (anchorage-independent growth) of highly invasive human breast cancer cells MDA-MB-231. GDNT suppressed expression of the cell cycle regulatory protein CDC20, which is over-expressed in precancerous and breast cancer cells compared to normal mammary epithelial cells. Moreover, we found that CDC20 is over-expressed in tumors when compared to the tissue surrounding the tumor in specimens from breast cancer patients. GDNT also inhibited invasive behavior (cell adhesion, cell migration, and cell invasion) through the suppression of secretion of urokinase-plasminogen activator (uPA) and inhibited expression of uPA receptor. In conclusion, mushroom GDNT is a natural agent that has potential as a therapy for invasive breast cancers.

  7. miR-483 is Down-Regulated in Polycystic Ovarian Syndrome and Inhibits KGN Cell Proliferation via Targeting Insulin-Like Growth Factor 1 (IGF1)

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    Xiang, Yungai; Song, Yuxia; Li, Yan; Zhao, Dongmei; Ma, Liying; Tan, Li

    2016-01-01

    Background Polycystic ovarian syndrome (PCOS) is a common metabolic disorder in premenopausal woman, characterized by hyperandrogenism, oligoanovulation, and insulin resistance. microRNAs play pivotal roles in regulating key factors of PCOS. However, relevant research remains limited. This study aimed to reveal the role and potential mechanism of miR-483 in PCOS. Material/Methods PCOS patients (n=20) were recruited for detecting miR-483 expression in lesion and normal ovary cortex. Human granulosa-like tumor cell line KGN was used to alter miR-483 expression by cell transfection. Cell viability and proliferation were analyzed by MTT assay and colony formation assay, and cell cycle was detected by flow cytometry. Interaction between miR-483 and IGF1 was verified by luciferase reporter assay. KGN cells were further treated by insulin to investigate the relationship between miR-483 and insulin. Results miR-483 was significantly down-regulated in lesion ovary cortex from PCOS patients (P<0.001). In KGN cells, overexpression of miR-483 inhibited cell viability and proliferation, and induced cell cycle arrest. miR-483 also inhibited CCNB1, CCND1, and CDK2. miR-483 sponge induced the opposite effects. miR-483 directly targeted IGF1 3′UTR, and IGF1 promoted KGN cell proliferation and reversed miR-483-inhibited cell viability. Insulin treatment in KGN cells inhibited miR-483, and promoted IGF1 and cell proliferation. Conclusions These results suggest that miR-483 is a PCOS suppressor inhibiting cell proliferation, possibly via targeting IGF1, and that it is involved in insulin-induced cell proliferation. miR-483 is a potential alternative for diagnosing and treating PCOS. PMID:27662007

  8. Curcumin inhibits cholesterol uptake in Caco-2 cells by down-regulation of NPC1L1 expression

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    Duan Rui-Dong

    2010-04-01

    Full Text Available Abstract Background Curcumin is a polyphenol and the one of the principle curcuminoids of the spice turmeric. Its antioxidant, anti-cancer and anti-inflammatory effects have been intensively studied. Previous in vivo studies showed that administration of curcumin also decreased cholesterol levels in the blood, and the effects were considered to be related to upregulation of LDL receptor. However, since plasma cholesterol levels are also influenced by the uptake of cholesterol in the gut, which is mediated by a specific transporter Niemann-Pick Cl-like 1 (NPC1L1 protein, the present study is to investigate whether curcumin affects cholesterol uptake in the intestinal Caco-2 cells. Methods Caco-2 cells were cultured to confluence. The micelles composed of bile salt, monoolein, and 14C-cholesterol were prepared. We first incubated the cells with the micelles in the presence and absence of ezetimibe, the specific inhibitor of NPC1L1, to see whether the uptake of the cholesterol in the cells was mediated by NPC1L1. We then pretreated the cells with curcumin at different concentrations for 24 h followed by examination of the changes of cholesterol uptake in these curcumin-treated cells. Finally we determined whether curcumin affects the expression of NPC1L1 by both Western blot analysis and qPCR quantification. Results We found that the uptake of radioactive cholesterol in Caco-2 cells was inhibited by ezetimibe in a dose-dependent manner. The results indicate that the uptake of cholesterol in this study was mediated by NPC1L1. We then pretreated the cells with 25-100 μM curcumin for 24 h and found that such a treatment dose-dependently inhibited cholesterol uptake with 40% inhibition obtained by 100 μM curcumin. In addition, we found that the curcumin-induced inhibition of cholesterol uptake was associated with significant decrease in the levels of NPC1L1 protein and NPC1L1 mRNA, as analyzed by Western blot and qPCR, respectively. Conclusion

  9. miR-10b*, a master inhibitor of the cell cycle, is down-regulated in human breast tumours.

    Science.gov (United States)

    Biagioni, Francesca; Bossel Ben-Moshe, Noa; Fontemaggi, Giulia; Canu, Valeria; Mori, Federica; Antoniani, Barbara; Di Benedetto, Anna; Santoro, Raffaela; Germoni, Sabrina; De Angelis, Fernanda; Cambria, Anna; Avraham, Roi; Grasso, Giuseppe; Strano, Sabrina; Muti, Paola; Mottolese, Marcella; Yarden, Yosef; Domany, Eytan; Blandino, Giovanni

    2012-11-01

    Deregulated proliferation is a hallmark of cancer cells. Here, we show that microRNA-10b* is a master regulator of breast cancer cell proliferation and is downregulated in tumoural samples versus matched peritumoural counterparts. Two canonical CpG islands (5 kb) upstream from the precursor sequence are hypermethylated in the analysed breast cancer tissues. Ectopic delivery of synthetic microRNA-10b* in breast cancer cell lines or into xenograft mouse breast tumours inhibits cell proliferation and impairs tumour growth in vivo, respectively. We identified and validated in vitro and in vivo three novel target mRNAs of miR-10b* (BUB1, PLK1 and CCNA2), which play a remarkable role in cell cycle regulation and whose high expression in breast cancer patients is associated with reduced disease-free survival, relapse-free survival and metastasis-free survival when compared to patients with low expression. This also suggests that restoration of microRNA-10b* expression might have therapeutic promise.

  10. miR-10b*, a master inhibitor of the cell cycle, is down-regulated in human breast tumours

    Science.gov (United States)

    Biagioni, Francesca; Bossel Ben-Moshe, Noa; Fontemaggi, Giulia; Canu, Valeria; Mori, Federica; Antoniani, Barbara; Di Benedetto, Anna; Santoro, Raffaela; Germoni, Sabrina; De Angelis, Fernanda; Cambria, Anna; Avraham, Roi; Grasso, Giuseppe; Strano, Sabrina; Muti, Paola; Mottolese, Marcella; Yarden, Yosef; Domany, Eytan; Blandino, Giovanni

    2012-01-01

    Deregulated proliferation is a hallmark of cancer cells. Here, we show that microRNA-10b* is a master regulator of breast cancer cell proliferation and is downregulated in tumoural samples versus matched peritumoural counterparts. Two canonical CpG islands (5 kb) upstream from the precursor sequence are hypermethylated in the analysed breast cancer tissues. Ectopic delivery of synthetic microRNA-10b* in breast cancer cell lines or into xenograft mouse breast tumours inhibits cell proliferation and impairs tumour growth in vivo, respectively. We identified and validated in vitro and in vivo three novel target mRNAs of miR-10b* (BUB1, PLK1 and CCNA2), which play a remarkable role in cell cycle regulation and whose high expression in breast cancer patients is associated with reduced disease-free survival, relapse-free survival and metastasis-free survival when compared to patients with low expression. This also suggests that restoration of microRNA-10b* expression might have therapeutic promise. PMID:23125021

  11. MiR-122 increases sensitivity of drug-resistant BEL-7402/5-FU cells to 5-fluorouracil via down-regulation of bcl-2 family proteins.

    Science.gov (United States)

    Yin, Jie; Tang, Hui Fang; Xiang, Qiong; Yu, Jia; Yang, Xiao Yang; Hu, Nan; Lei, Xiao Yong

    2011-12-01

    To investigate the changes in drug sensitivity of miR-122 transfected BEL-7402/5-FU cells. MiR-122 and negative miRNA expression vectors were constructed and stably transfected into BEL-7402/5-FU cells. Real-time RT-PCR was used to detect the level of miR-122, Bcl-XL, Bcl-2 and P53 mRNA. Western Blotting was used to detect Bcl-2, Bcl-XL and P53 protein expression. Drug sensitivity of the cells to 5-fluorouracil (5-FU) was analyzed with MTT and flow cytometry. Compared with negative miRNA transfectants or untreated cells, mRNA and protein expression level of Bcl-2, Bcl-XL in stable miR-122 transfectants were decreased. Accordingly, P53 protein expression showed a significant up-regulation; MTT results showed that after incubation with 5-FU, miR-122 transfectants had higher cell inhibitory rates than negative miRNA or untreated cells; flow cytometry results demonstrated that apoptosis rate increased in miR-122 transfected cells, compared with negative miRNA or untreated cells. After addition of 5-FU (10 and 100 micromol/I), miR-122 transfected cells showed higher apoptosis rate than negative miRNA or untreated cells. MiR-122 can specifically down-regulate the expression of Bcl-2 and Bcl-XL, and increase P53 activity in BEL-7402/5-FU cells, which increased cells spontaneous apoptosis and sensitize cells to 5-FU. Therefore, MiR-122 can be used as a potential therapy agent against human hepatoblastoma.

  12. Juglone, isolated from Juglans mandshurica Maxim, induces apoptosis via down-regulation of AR expression in human prostate cancer LNCaP cells.

    Science.gov (United States)

    Xu, Huali; Yu, Xiaofeng; Qu, Shaochun; Sui, Dayun

    2013-06-15

    Juglone is a natural compound which has been isolated from Juglans mandshurica Maxim. Recent studies have shown that juglone had various pharmacological effects such as anti-viral, anti-bacterial and anti-cancer. However, its anti-cancer activity on human prostate cancer LNCaP cell has not been examined. Thus, the current study was designed to elucidate the molecular mechanism of apoptosis induced by juglone in androgen-sensitive prostate cancer LNCaP cells. MTT assay was performed to examine the anti-proliferative effect of juglone. Occurrence of apoptosis was detected by Hoechst 33342 staining and flow cytometry in LNCaP cells treated with juglone for 24h. The result shown that juglone inhibited the growth of LNCaP cells in a dose-dependent manner. Morphological changes of apoptotic body formation after juglone treatment were observed by Hoechst 33342 staining. This apoptotic induction was associated with loss of mitochondrial membrane potential, and caspase-3, -9 activation. Moreover, we found that juglone significantly inhibited the expression levels of androgen receptor (AR) and prostate-specific antigen (PSA) in a dose-dependent manner, as well as abrogated up-regulation of AR and PSA genes with and/or without dihydrotestosterone (DHT). Take together, our results demonstrated that juglone might induce the apoptosis in LNCaP cell via down-regulation of AR expression. Therefore, our results indicated that juglone may be a potential candidate of drug for androgen-sensitive prostate cancer.

  13. Nrf2 is crucial for the down-regulation of Cyp7a1 induced by arachidonic acid in Hepg2 cells.

    Science.gov (United States)

    Zhang, Jin-Ming; Wang, Xing-He; Hao, Li-Hong; Wang, He; Zhang, Xiu-Ying; Muhammad, Ishfaq; Qi, Yue; Li, Guang-Liang; Sun, Xiao-Qi

    2017-03-07

    In former research, cyp7a1 expression was decreased but Nrf2 transcription and hepatic arachidonic acid (AA) concentration were increased in high-fat diet fed mice. This study aims to investigate the influence of AA in CYP7A1 expression and the role of Nrf2 in regulating CYP7A1 in the process. HepG2 cells were administered with different concentrations of AA. Nrf2 and CYP7A1 expressions were analyzed by real-time PCR and western blot. Nrf2 silenced and over-expressed cell models were constructed by Nrf2 siRNA and eukaryotic expression vector transient transfections and were used to investigate the role of Nrf2 in regulating CYP7A1 following AA administration. The results showed that Nrf2 was increased dose-dependently but CYP7A1 was decreased dose-dependently in cells treated with increasing concentrations of AA. The expression of CYP7A1 was increased by Nrf2 silence and was decreased by Nrf2 over-expression in HepG2 cells treated with different concentrations of AA. In conclusion, Nrf2 plays a significant role in the down-regulation of CYP7A1 induced by AA in HepG2 cells.

  14. Cordycepin Down-Regulates Multiple Drug Resistant (MDR/HIF-1α through Regulating AMPK/mTORC1 Signaling in GBC-SD Gallbladder Cancer Cells

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    Wei-Ding Wu

    2014-07-01

    Full Text Available Gallbladder cancer is the most common malignancy of the bile duct, with low 5-year survival rate and poor prognosis. Novel effective treatments are urgently needed for the therapy of this disease. Here, we showed that cordycepin, the bioactive compound in genus Cordyceps, induced growth inhibition and apoptosis in cultured gallbladder cancer cells (Mz-ChA-1, QBC939 and GBC-SD lines. We found that cordycepin inhibited mTOR complex 1 (mTORC1 activation and down-regulated multiple drug resistant (MDR/hypoxia-inducible factor 1α (HIF-1α expression through activating of AMP-activated protein kinase (AMPK signaling in gallbladder cancer GBC-SD cells. Contrarily, AMPKα1-shRNA depletion dramatically inhibited cordycepin-induced molecular changes as well as GBC-SD cell apoptosis. Further, our results showed that co-treatment with a low concentration cordycepin could remarkably enhance the chemosensitivity of GBC-SD cells to gemcitabine and 5-fluorouracil (5-FU, and the mechanism may be attributed to AMPK activation and MDR degradation. In summary, cordycepin induces growth inhibition and apoptosis in gallbladder cancer cells via activating AMPK signaling. Cordycepin could be a promising new drug or chemo-adjuvant for gallbladder cancer.

  15. Regulated Production of Mature Insulin in Rat Hepatoma Cells:Insulin Production is Up-regulated by Dexamethasone and Down-regulated by Insulin

    Institute of Scientific and Technical Information of China (English)

    Xin-Yu QIN; Kun-Tang SHEN; Lu-Jun SONG; Xin ZHANG; Ze-Guang HAN

    2006-01-01

    We engineered an artificial β cell line that produces an up-regulation of insulin in response to dexamethasone, and a down-regulation in response to insulin. A regulatory secretion system was devised by placing proinsulin cDNA containing genetically engineered furin endoprotease cleavage sites and a regulatory promoter for phosphoenolpyruvate carboxykinase (PEPCK), and an insulin expressing retrovirus vector (pN-PEPCK-mINS) was constructed and transfected into Hepa1-6 cells. The levels of insulin in culture medium and expression of insulin gene was estimated by radioimmunoassay and reverse transcriptionpolymerase chain reaction (RT-PCR), respectively. The clone (Hepa1-6/INS21), which secreted the highest level of insulin (10.79 μIU/106 cells per day), was selected for the regulation experiment. Compared with the non-treated Hepa1-6/INS21 cells, insulin production was augmented 3.6-fold by the addition of 10-7 M of dexamethasone. Addition of exogenous insulin to the culture medium decreased insulin mRNA expression remarkably on RT-PCR results, while dexamethasone increased insulin gene expression at the transcriptional level. The data indicated that genetically engineered Hepa1-6 cells could synthesize process and secrete insulin in a physiological manner.

  16. T-helper Cell Type-1 Transcription Factor T-Bet Is Down-regulated in Type 1 Diabetes

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    Hajar Vaseghi

    2016-10-01

    Full Text Available T cells have been identified as key players in the pathogenesis of type 1 diabetes. However, the exact role of T-cell subpopulations in this pathway is presently unknown. The purpose of this study was to assess the expression pattern of two lineage-specifying transcription factors GATA-3 and T-bet, which are important in T helper type 1 (Th1 and Th2 cell development, respectively. Gene expression analysis of peripheral blood mononuclear cells (PBMCs was performed using reverse transcription-quantitative polymerase chain reaction (RT-qPCR. Plasma levels of IFN-γ and IL-4 were also determined by ELISA. T-bet and IFN-γ gene expression was significantly lower in patients group compared with healthy controls (p<0.05. The expression of GATA-3 was relatively similar in patients and controls; however, IL-4 mRNAs were significantly increased in the PBMCs from patients as compared with normal controls (p<0.05. In addition, a marked increase in plasma IL-4 levels were observed in patient group compared with controls (p<0.001. To the contrary, IFN-γ protein levels were decreased in patients in comparison with controls (p<0.001. These data suggest additional implications of the role of Th1/Th2 imbalance for the immunopathogenesis of type 1 diabetes. 

  17. Induction of differentiation by down-regulation of Nanog and Rex-1 in cord blood derived unrestricted somatic stem cells.

    Science.gov (United States)

    Langroudi, Lida; Forouzandeh, Mehdi; Soleimani, Masoud; Atashi, Amir; Golestaneh, Azadeh Fahim

    2013-07-01

    Stem cells with high self-renewal and tissue regeneration potentials are the core components of regenerative medicine. Adult stem cells with many available sources, high repairing ability, and also possessing no ethical issues are popular candidates in the clinical field. In this study we looked upon the effects of two transcription factors Nanog and Rex-1 in self-renewal and differentiation abilities of a subpopulation of cord blood stem cells known as unrestricted somatic stem cells (USSCs). USSCs were expanded and transfected in vitro with siRNAs targeting either Nanog, Rex-1, and in combination. Gene suppressions were achieved at both transcript and proteome level. Differentiations were evaluated by specific Real time PCR and differentiating staining. Nanog knock down revealed a significant increase in osteogenic markers, Osteocalcin and Osteopontin expression as well as a positive Alizarin Red staining, which proposes Osteogenesis. This treatment also became positive for Oil Red staining, implying adipogenic differentiation as well. In contrast, Rex-1 knock down showed an increase in MAP II and Nestin expression, which is a hall mark of neural differentiation. Surprisingly, treatment with both siRNAs did not express any changes in any of the assessed markers. Therefore, our results indicated a bilateral mesenchymal differentiation for Nanog and a neural lineage fate for Rex-1 suppression. Considering that both transcription factors are core activators of self-renewal and also are orchestrating with other factors, our results imply a positive feedback in response to changes in the regulatory network of self-renewal.

  18. The Cytoplasmic Domain of CD4 Is Sufficient for Its Down-Regulation from the Cell Surface by Human Immunodeficiency Virus Type 1 Nef

    OpenAIRE

    S. J. Anderson; Lenburg, M; Landau, N R; Garcia, J. V.

    1994-01-01

    Human immunodeficiency virus type 1 Nef down-regulates surface expression of murine and human CD4 but not human CD8. We recently reported that the cytoplasmic domain of CD4 is required for its down-regulation by Nef. Using a chimeric molecule composed of the extracellular and transmembrane domains of human CD8 fused to the cytoplasmic domain of human CD4, we show here that the cytoplasmic domain of CD4 is sufficient for down-regulation by Nef. Since the cytoplasmic domain of CD4 is also the s...

  19. Periostin down-regulation attenuates the pro-fibrogenic response of hepatic stellate cells induced by TGF-β1.

    Science.gov (United States)

    Hong, Li; Shejiao, Dai; Fenrong, Chen; Gang, Zhao; Lei, Dong

    2015-10-01

    Liver fibrosis is characterized by an exacerbated accumulation of deposition of the extracellular matrix (ECM), and the activation of hepatic stellate cells (HSC) plays a pivotal role in the development of liver fibrosis. Periostin has been shown to regulate cell adhesion, proliferation, migration and apoptosis; however, the involvement of periostin and its role in transforming growth factor (TGF)-β1-induced HSC activation remains unclear. We used RT-PCR and Western blot to evaluate the expression level of periostin in hepatic fibrosis tissues and HSCs, respectively. Cell proliferation was determined using the Cell Proliferation ELISA BrdU kit, cell cycle was analysed by flow cytometry. The expression of α-smooth muscle actin (α-SMA), collagen I, TGF-β1, p-Smad2 and p-Smad3 were determined by western blot. Our study found that periostin was up-regulated in liver fibrotic tissues and activated HSCs. In addition, siRNA-periostin suppressed TGF-β1-induced HSC proliferation. The HSC transfected with siRNA-periostin significantly inhibited TGF-β1-induced expression levels of α-SMA and collagen I. Furthermore, TGF-β1 stimulated the expression of periostin, and siRNA-periostin attenuated TGF-β1-induced Smad2/3 activation in HSCs. These results suggest that periostin may function as a novel regulator to modulate HSC activation, potentially by promoting the TGF-β1/Smad signalling pathway, and propose a strategy to target periostin for the treatment of liver fibrosis.

  20. Down-regulating cyclin-dependent kinase 9 of alloreactive CD4+ T cells prolongs allograft survival

    Science.gov (United States)

    Zhan, Yang; Han, Yeming; Sun, Hukui; Liang, Ting; Zhang, Chao; Song, Jing; Hou, Guihua

    2016-01-01

    CDK9 (Cyclin-dependent kinase 9)/Cyclin T1/RNA polymerase II pathway has been demonstrated to promote the development of several inflammatory diseases, such as arthritis or atherosclerosis, however, its roles in allotransplantation rejection have not been addressed. Here, we found that CDK9/Cyclin T1 were apparently up-regulated in the allogeneic group, which was positively correlated with allograft damage. CDK9 was inhibited obviously in naive splenic CD4+ T cells treated 6 h with 3 μM PHA767491 (a CDK9 inhibitor), and adoptive transfer of these CD4+ T cells into allografted SCID mice resulted in prolonged survival compared with the group without PHA767491 pretreated. Decelerated rejection was correlated with enhanced IL-4 and IL-10 production and with decreased IFN-γ production by alloreactive T cells. More interestingly, we found that CDK942, not CDK955, was high expressed in allorejection group, which could be prominently dampened with PHA767491 treatment. The expression of CDK942 was consistent with its downstream molecule RNA polymerase II. Altogether, our findings revealed the crucial role of CDK9/Cyclin T1/Pol II pathway in promoting allorejection at multiple levels and may provide a new approach for transplantation tolerance induction through targeting CDK9. PMID:27102157

  1. Mechanisms controlling the smooth muscle cell death in progeria via down-regulation of poly(ADP-ribose) polymerase 1.

    Science.gov (United States)

    Zhang, Haoyue; Xiong, Zheng-Mei; Cao, Kan

    2014-06-03

    Hutchinson-Gilford progeria syndrome (HGPS) is a severe human premature aging disorder caused by a lamin A mutant named progerin. Death occurs at a mean age of 13 y from cardiovascular problems. Previous studies revealed loss of vascular smooth muscle cells (SMCs) in the media of large arteries in a patient with HGPS and two mouse models, suggesting a causal connection between the SMC loss and cardiovascular malfunction. However, the mechanisms of how progerin leads to massive SMC loss are unknown. In this study, using SMCs differentiated from HGPS induced pluripotent stem cells, we show that HGPS SMCs exhibit a profound proliferative defect, which is primarily caused by caspase-independent cell death. Importantly, progerin accumulation stimulates a powerful suppression of PARP1 and consequently triggers an activation of the error-prone nonhomologous end joining response. As a result, most HGPS SMCs exhibit prolonged mitosis and die of mitotic catastrophe. This study demonstrates a critical role of PARP1 in mediating SMC loss in patients with HGPS and elucidates a molecular pathway underlying the progressive SMC loss in progeria.

  2. Doxycycline down-regulates DNA-PK and radiosensitizes tumor initiating cells: Implications for more effective radiation therapy.

    Science.gov (United States)

    Lamb, Rebecca; Fiorillo, Marco; Chadwick, Amy; Ozsvari, Bela; Reeves, Kimberly J; Smith, Duncan L; Clarke, Robert B; Howell, Sacha J; Cappello, Anna Rita; Martinez-Outschoorn, Ubaldo E; Peiris-Pagès, Maria; Sotgia, Federica; Lisanti, Michael P

    2015-06-10

    DNA-PK is an enzyme that is required for proper DNA-repair and is thought to confer radio-resistance in cancer cells. As a consequence, it is a high-profile validated target for new pharmaceutical development. However, no FDA-approved DNA-PK inhibitors have emerged, despite many years of drug discovery and lead optimization. This is largely because existing DNA-PK inhibitors suffer from poor pharmacokinetics. They are not well absorbed and/or are unstable, with a short plasma half-life. Here, we identified the first FDA-approved DNA-PK inhibitor by "chemical proteomics". In an effort to understand how doxycycline targets cancer stem-like cells (CSCs), we serendipitously discovered that doxycycline reduces DNA-PK protein expression by nearly 15-fold (> 90%). In accordance with these observations, we show that doxycycline functionally radio-sensitizes breast CSCs, by up to 4.5-fold. Moreover, we demonstrate that DNA-PK is highly over-expressed in both MCF7- and T47D-derived mammospheres. Interestingly, genetic or pharmacological inhibition of DNA-PK in MCF7 cells is sufficient to functionally block mammosphere formation. Thus, it appears that active DNA-repair is required for the clonal expansion of CSCs. Mechanistically, doxycycline treatment dramatically reduced the oxidative mitochondrial capacity and the glycolytic activity of cancer cells, consistent with previous studies linking DNA-PK expression to the proper maintenance of mitochondrial DNA integrity and copy number. Using a luciferase-based assay, we observed that doxycycline treatment quantitatively reduces the anti-oxidant response (NRF1/2) and effectively blocks signaling along multiple independent pathways normally associated with stem cells, including STAT1/3, Sonic Hedgehog (Shh), Notch, WNT and TGF-beta signaling. In conclusion, we propose that the efficacy of doxycycline as a DNA-PK inhibitor should be tested in Phase-II clinical trials, in combination with radio-therapy. Doxycycline has excellent

  3. Down-regulation of IL-8 expression in human airway epithelial cells through helper-dependent adenoviral-mediated RNA interference

    Institute of Scientific and Technical Information of China (English)

    Huibi CAO; Anan WANG; Bernard MARTIN; David R.KOEHLER; Pamela L.ZEITLIN; A.Keith TANAWELL; Jim HU

    2005-01-01

    Interleukin (IL)-8 is a potent neutrophil chemotactic factor and a crucial mediator in neutrophil-dependent inflammation.Various cell types produce IL-8, either in response to external stimuli such as cytokines or bacterial infection, or after malignant transformation. Anti-IL-8 strategies have been considered for anti-inflammatory therapy. In this paper we demonstrate that the RNA interference technique can be used to efficiently down-regulate IL-8 protein expression in airway epithelial cells. We used a helper-dependent adenoviral vector to express a small hairpin (sh)RNA targeting human IL-8 in cultured airway epithelial cells (IB3-1, Cftr-/-; C38, Cftr-corrected) stimulated with TNF-α, IL-1 β or heat-inactivated Burkholderia cenocepacia. Stimulated IL-8 expression in IB3-1 and C38 cells was significantly reduced by shRNA expression. The shRNA targeting IL-8 had no effect on the activation of NF-κB, or on the protein levels of Iκ B or IL-6, suggesting that this anti-IL-8 strategy was highly specific, and therefore may offer potential for the treatment of inflammatory diseases.

  4. Effect of NS398 on inducing apoptosis and down-regulating Bcl-2 protein in human osteosarcoma cell MG-63 line

    Institute of Scientific and Technical Information of China (English)

    Eryou Feng; Biao Gao; Renyun Xia

    2005-01-01

    Objective: This study investigated the effect of NS398 on anti-proliferation and the mechanism of inducing apoptosis of human osteosarcoma cell MG-63 line in vitro. Methods: Growth suppression was detected by MTT method. Special morphological changes of apoptosis were observed by transmission electron microscopy (TEM) and DNA fragments electrophoresis. The apoptotic rates were quantified by flow cytometry (FCM). And Bcl-2 protein level were detected by Western Blotting assay. Results: Different concentration NS398 inhibited the cell growth. Through TEM and DNA electrophoresis, the special morphological changes were observed after 24,48 and 72 h treatment with NS398. The apoptotic rates of MG-63 cells treated with NS398 were respectively, significantly higher than that of the control group( P < 0.01). Western blot assay showed that NS398 reduced Bcl-2 protein expression. Conclusion: NS398 suppressed the proliferation of human osteosarcoma cell MG-63 line and induced apoptosis. The mechanism may be associated with down-regulation expression level of bcl-2 protein.

  5. Targeted DNA Methylation by a DNA Methyltransferase Coupled to a Triple Helix Forming Oligonucleotide To Down-Regulate the Epithelial Cell Adhesion Molecule

    NARCIS (Netherlands)

    van der Gun, Bernardina T. F.; Maluszynska-Hoffman, Maria; Kiss, Antal; Arendzen, Alice J.; Ruiters, Marcel H. J.; McLaughlin, Pamela M. J.; Weinhold, Elmar; Rots, Marianne G.

    2010-01-01

    The epithelial cell adhesion molecule (EpCAM) is a membrane glycoprotein that has been identified as a marker of cancer-initiating cells. EpCAM is highly expressed on most carcinomas, and transient silencing of EpCAM expression leads to reduced oncogenic potential. To silence (he EpCAM gene in a per

  6. Chronic Hyperinsulinism Induced Down-regulation of Insulin Post-Recentor Signaling Transduction in Hep G2 Cells

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Summary: To study the regulatory effect of acute and chronic insulin treatment on insulin post-re-ceptor signaling transduction pathway in a human hepatoma cell line (Hep G2), Hep G2 cells wereincubated in the presence or absence of insulin with different concentrations in serum free mediafor 16 h and then stimulated with 100 nmol/L insulin for 1 min. Protein levels of insulin receptorβ-subunit (IRβ), insulin receptor substrate-1 (IRS-1) and p85 subunit of phosphatidylinositol 3-kinase (PI 3-kinase) were determined in total cell lysates by Western-immunoblot. Phosphorylat-ed proteins IRβ, IRS-1 and interaction of PI 3-kinase with IRS-1 were determined by immunopre-cipitation. Results showed that 1-min insulin stimulation rapidly induced tyrosine phosphorylationof IRβ and IRS-l, which in turn, resulting in association of PI 3-kinase with IRS-1. 1-100 nmol/L chronic insulin treatment induced a dose-dependent decrease in the protein level of IRβ and aslight decrease in the protein level of IRS-1. There wass more marked reduction in the phospho-rylation of IRβ, IRS-1, reaching a nadir of 22 % (P<0. 01) and 15 % (P<0. 01) of control lev-els, respectively, after 16 h treatment with 100 nmol/L insulin. The association between IRS-1and PI 3-kinase was decreased by 66 % (P<0. 01). There was no significant change in PI 3-ki-nase protein levels. These data suggest that chronic insulin treatment can induce alterations ofIRβ, IRS-1 and PI 3-kinase three early steps in insulin action, which contributes significantly toinsulin resistance, and may account for desensitization of insulin action.

  7. DOWN-REGULATION OF INDUCIBLE NITRIC OXIDE SYNTHASE EXPRESSION BY INOSITOL HEXAPHOSPHATE IN HUMAN COLON CANCER CELLS.

    Science.gov (United States)

    Kapral, Małgorzata; Wawszczyk, Joanna; Sośnicki, Stanisław; Węglarz, Ludmiła

    2015-01-01

    Inflammatory bowel disease (IBD) is chronic inflammatory condition associated with increased risk of developing colorectal cancer. A number of mediators of inflammation, such as pro-inflammatory cytokines, prostaglandins and nitric oxide have been involved in carcinogenesis, especially in the promotion and progression stages. NO is synthesized from L-arginine by constitutively expressed endothelial and neuronal nitric oxide synthases (eNOS and nNOS, respectively) and an inducible NOS (iNOS) isoform expressed under inflammatory conditions. A selective inhibitors of iNOS could be, therefore, considered to be good candidates as chemopreventive agents against colon cancer. In this study, the effect of inositol hexaphosphate (IP6), dietary phytochemical, on the mRNA expression of iNOS stimulated with bacterial lipopolysaccharides (Escherichia coli and Salmonella typhimurium) and IL-1β in intestinal cells Caco-2 for 6 and 12 h was investigated. A transcription level of iNOS with the use real time QRT-PCR technique was determined in cells treated with 1 and 2.5 mM IP6. Stimulation of Caco-2 with pro-inflammatory factors (LPS and IL-1β) resulted in an up-expression of iNOS mRNA at 6 and 12 h. Cells exposed to IP6 only revealed significant reduction in iNOS gene transcription after 12 h. A decrease in iNOS transcription by IP6 following the gene induction by proinflammatory agents in 6 and 12 h lasting cultures was also determined. The findings of this study suggest that one of the anti-cancer and anti-inflammatory abilities of IP6 can be realized by suppressing the expression of gene encoding inducible nitric oxide synthase isoform at the transcriptional level.

  8. Alpha-2-glycoprotein 1(AZGP1 regulates biological behaviors of LoVo cells by down-regulating mTOR signaling pathway and endogenous fatty acid synthesis.

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    Ligong Chang

    Full Text Available AZGP1 is a multifaceted protein associated with lipid mobilization, a process that is regulated by FASN and other metabolic pathways such as mTOR signaling. The active mTOR signaling pathway has been found to be involved in a variety of tumors. However, it remains unclear whether it is involved in the regulation of AZGP1 and FASN. An AZGP1-expressing plasmid was transfected into a human colorectal cancer cell line (LoVo with a low expression of AZGP1. The expression of AZGP1, FASN, eIF4E, p-mTOR, p-S6,and S6K1 were measured by Western blot analysis, and target genes were detected by RT-PCR. Cell proliferation was studied using the MTT and colony formation assays. The analysis of apoptosis and the cell cycle phase were assessed by flow cytometry. The capacity of cell migration was investigated using the transwell migration assay. We found that the expression of AZGP1 was up-regulated while the expression of FASN, eIF4E, p-mTOR, p-S6, and S6K1 were down-regulated in LoVo cells after AZGP1 was expressed. The proliferation of malignant cells was reduced in AZGP1-overexpression cells, which is consistent with an increased in the G2-arrest and apoptosis rate. Furthermore, the migration of AZGP1-overexpression cells was decreased. The overexpression of AZGP1 suppressed the activation of the mTOR pathway and endogenous FASN-regulated fatty acid synthesis, mitigating the malignant phenotype of LoVo cells. Herein, we provide evidence that AZGP1 may constitute a novel tumor suppressor for LoVo colorectal cancer cells.

  9. Alpha-2-glycoprotein 1(AZGP1) regulates biological behaviors of LoVo cells by down-regulating mTOR signaling pathway and endogenous fatty acid synthesis.

    Science.gov (United States)

    Chang, Ligong; Tian, Xiaoqiang; Lu, Yinghui; Jia, Min; Wu, Peng; Huang, Peilin

    2014-01-01

    AZGP1 is a multifaceted protein associated with lipid mobilization, a process that is regulated by FASN and other metabolic pathways such as mTOR signaling. The active mTOR signaling pathway has been found to be involved in a variety of tumors. However, it remains unclear whether it is involved in the regulation of AZGP1 and FASN. An AZGP1-expressing plasmid was transfected into a human colorectal cancer cell line (LoVo) with a low expression of AZGP1. The expression of AZGP1, FASN, eIF4E, p-mTOR, p-S6,and S6K1 were measured by Western blot analysis, and target genes were detected by RT-PCR. Cell proliferation was studied using the MTT and colony formation assays. The analysis of apoptosis and the cell cycle phase were assessed by flow cytometry. The capacity of cell migration was investigated using the transwell migration assay. We found that the expression of AZGP1 was up-regulated while the expression of FASN, eIF4E, p-mTOR, p-S6, and S6K1 were down-regulated in LoVo cells after AZGP1 was expressed. The proliferation of malignant cells was reduced in AZGP1-overexpression cells, which is consistent with an increased in the G2-arrest and apoptosis rate. Furthermore, the migration of AZGP1-overexpression cells was decreased. The overexpression of AZGP1 suppressed the activation of the mTOR pathway and endogenous FASN-regulated fatty acid synthesis, mitigating the malignant phenotype of LoVo cells. Herein, we provide evidence that AZGP1 may constitute a novel tumor suppressor for LoVo colorectal cancer cells.

  10. MiR674 inhibits the neuraminidase-stimulated immune response on dendritic cells via down-regulated Mbnl3.

    Science.gov (United States)

    Lin, Jian; Chen, Ya T; Xia, Jing; Yang, Qian

    2016-08-02

    Neuraminidase (NA), a structural protein of the H9N2 avian influenza virus (H9N2 AIV), can facilitate viral invasion of the upper airway by cleaving the sialic acid moieties on mucin. Dendritic cells (DCs) are major antigen-presenting cells whose immune functions, such as presenting antigens and activating lymphocytes, can be regulated by microRNAs. Here, we studied the molecular mechanism of miRNA-induced repression of immune responses in mouse DCs. First, we screened for and verified the miRNAs induced by NA. Then, we showed that, consistent with the H9N2 virus treatment, the viral NA up-regulated the expression of miR-155, miR-674, and miR-499 in DCs; however, unlike H9N2 virus treatment, the presence of NA was associated with reduced expression of miR-181b1. Our results suggest that NA significantly increased DC surface markers CD80 and MHCII and enhanced the ability of activating lymphocytes and secreting cytokines compared with HA, NP and M2. Meanwhile, we found that miR-674 and miR-155 over-expression increased all surface markers of DC. Nevertheless, by inhibiting the expression of miR-674 and miR-155, NA lost the ability to promote DC maturation. Furthermore, we predicted and demonstrated that Pgm2l1, Aldh18a1, Camk1d, and Mbnl3 were the target genes of miR-674. Among them, Mbnl3 interference strongly blocked the mature DCs. Collectively, our data shed new light on the roles of and mechanisms involved in the repression of DCs by miRNAs, which may contribute to efforts to develop a prophylaxis for the influenza virus.

  11. Vitamin C down-regulate apo(a) expression via Tet2-dependent DNA demethylation in HepG2 cells.

    Science.gov (United States)

    Qu, Kai; Ma, Xiao-Feng; Li, Guo-Hua; Zhang, Hai; Liu, Ya-Mi; Zhang, Kai; Zeng, Jun-Fa; Lei, Jian-Jun; Wei, Dang-Heng; Wang, Zuo

    2017-05-01

    Lipoprotein(a)[Lp(a)] is a risk factor for coronary heart diseases. However, the metabolism of this protein remains poorly understood. Efficient and specific drugs that can decrease high plasma levels of Lp(a) have not been developed yet. Vitamin C is responsible for maintaining the catalytic activity of a group of iron and 2-oxoglutarate (2OG)-dependent dioxygenases and induces the generation of 5-hydroxymethylcytosine (5hmC) via Ten-eleven translocation (Tet) dioxygenases. In addition, It has been reported vitamin C deficiency induces atherosclerosis and increases Lp(a) and apo(a) plasma levels in Lp(a)+ mice. However, the mechanism is still unclear. In this study, we investigated the effects of vitamin C on apo(a) expression and the possible molecular mechanism of vitamin C that influences apolipoprotein(a) [apo(a)] biosynthesis in HepG2 cells. Results showed that vitamin C significantly inhibited the expression and secretion levels of apo(a). Vitamin C can also increase ELK1 expression and hydroxymethylation of ELK1 promoter and the globle DNA in HepG2 cells. In addition, the effects of vitamin C inhibiting the apo(a) expression were attenuated by ELK1siRNA and Tet2siRNA. These results suggested vitamin C down-regulate apo(a) expression via Tet2-dependent DNA demethylation in HepG2 cells.

  12. TGF-β1 treated murine dendritic cells are maturation resistant and down-regulate Toll-like receptor 4 expression

    Institute of Scientific and Technical Information of China (English)

    牟海波; 林茂芳; 岑洪; 俞静; 孟筱坚

    2004-01-01

    Objective: To explore theeffects of transforming growth factor β1 (TGF-β1) on dendritic cells (DC). Methods:Murine bone marrow cells were cultured with GM-CSF and TGF-β1 to develop TGF-β1-treated DC (TGFβ-DC). Then they were stimulated by lipopolysaccharide (LPS). Their phenotypes were assessed by flow cytometry (FCM). The allogeneic stimulating capacity of DC was measured by mixed lymphocyte reaction (MLR) using BrdU ELISA method and IL-12p70protein was detected by ELISA. The expression of Toil-like receptor 4 (TLR4) was analyzed by semi quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and FCM. Results: Compared to immature DC (imDC) cultured by GM-CSF alone, the TGFβ-DC express lower CD80, CD86, I-Ab and CD40. The TGFβ-DC were resistant to maturation with LPS. Maturation resistance was evident from a failure to up-regulate co-stimulatory molecules (CMs), to stimulate larger T cells proliferation and to enhance secretion of IL-12p70. We also found that TGF-β 1 could down-regulate TLR4 expression on TGFβ-DC. Conclusion: TGFβ-DC are resistant to maturation stimulus (LPS) and might have some correlation with the down-modulation of TLR4 expression.

  13. Down-Regulation of Ca2+-Activated K+ Channel KCa1.1 in Human Breast Cancer MDA-MB-453 Cells Treated with Vitamin D Receptor Agonists

    Directory of Open Access Journals (Sweden)

    Anowara Khatun

    2016-12-01

    Full Text Available Vitamin D (VD reduces the risk of breast cancer and improves disease prognoses. Potential VD analogs are being developed as therapeutic agents for breast cancer treatments. The large-conductance Ca2+-activated K+ channel KCa1.1 regulates intracellular Ca2+ signaling pathways and is associated with high grade tumors and poor prognoses. In the present study, we examined the effects of treatments with VD receptor (VDR agonists on the expression and activity of KCa1.1 in human breast cancer MDA-MB-453 cells using real-time PCR, Western blotting, flow cytometry, and voltage-sensitive dye imaging. Treatments with VDR agonists for 72 h markedly decreased the expression levels of KCa1.1 transcripts and proteins in MDA-MB-453 cells, resulting in the significant inhibition of depolarization responses induced by paxilline, a specific KCa1.1 blocker. The specific proteasome inhibitor MG132 suppressed VDR agonist-induced decreases in KCa1.1 protein expression. These results suggest that KCa1.1 is a new downstream target of VDR signaling and the down-regulation of KCa1.1 through the transcriptional repression of KCa1.1 and enhancement of KCa1.1 protein degradation contribute, at least partly, to the antiproliferative effects of VDR agonists in breast cancer cells.

  14. A viral microRNA down-regulates multiple cell cycle genes through mRNA 5'UTRs.

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    Finn Grey

    Full Text Available Global gene expression data combined with bioinformatic analysis provides strong evidence that mammalian miRNAs mediate repression of gene expression primarily through binding sites within the 3' untranslated region (UTR. Using RNA induced silencing complex immunoprecipitation (RISC-IP techniques we have identified multiple cellular targets for a human cytomegalovirus (HCMV miRNA, miR-US25-1. Strikingly, this miRNA binds target sites primarily within 5'UTRs, mediating significant reduction in gene expression. Intriguingly, many of the genes targeted by miR-US25-1 are associated with cell cycle control, including cyclin E2, BRCC3, EID1, MAPRE2, and CD147, suggesting that miR-US25-1 is targeting genes within a related pathway. Deletion of miR-US25-1 from HCMV results in over expression of cyclin E2 in the context of viral infection. Our studies demonstrate that a viral miRNA mediates translational repression of multiple cellular genes by targeting mRNA 5'UTRs.

  15. Inhibition of p70S6K2 down-regulates Hedgehog/GLI pathway in non-small cell lung cancer cell lines

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    Kotani Hidehito

    2009-07-01

    Full Text Available Abstract Background The Hedgehog (HH pathway promotes tumorigenesis in a diversity of cancers. Activation of the HH signaling pathway is caused by overexpression of HH ligands or mutations in the components of the HH/GLI1 cascade, which lead to increased transactivation of GLI transcription factors. Although negative kinase regulators that antagonize the activity of GLI transcription factors have been reported, including GSK3β, PKA and CK1s, little is known regarding positive kinase regulators that are suitable for use on cancer therapeutic targets. The present study attempted to identify kinases whose silencing inhibits HH/GLI signalling in non-small cell lung cancer (NSCLC. Results To find positive kinase regulators in the HH pathway, kinome-wide siRNA screening was performed in a NSCLC cell line, A549, harboring the GLI regulatory reporter gene. This showed that p70S6K2-silencing remarkably reduced GLI reporter gene activity. The decrease in the activity of the HH pathway caused by p70S6K2-inhibition was accompanied by significant reduction in cell viability. We next investigated the mechanism for p70S6K2-mediated inhibition of GLI1 transcription by hypothesizing that GSK3β, a negative regulator of the HH pathway, is activated upon p70S6K2-silencing. We found that phosphorylated-GSK3β (Ser9 was reduced by p70S6K2-silencing, causing a decreased level of GLI1 protein. Finally, to further confirm the involvement of p70S6K2 in GLI1 signaling, down-regulation in GLI-mediated transcription by PI3KCA-inhibition was confirmed, establishing the pivotal role of the PI3K/p70S6K2 pathway in GLI1 cascade regulation. Conclusion We report herein that inhibition of p70S6K2, known as a downstream effector of the PI3K pathway, remarkably decreases GLI-mediated transactivation in NSCLC by reducing phosphorylated-GSK3β followed by GLI1 degradation. These results infer that p70S6K2 is a potential therapeutic target for NSCLC with hyperactivated HH/GLI pathway.

  16. [Down-regulated βIII-tubulin expression can reverse paclitaxel resistance in A549/taxol cells lines].

    Science.gov (United States)

    Zhuo, Yinling; Guo, Qisen

    2014-08-20

    背景与目的 化疗耐药导致肿瘤很快复发和/或转移,是目前肺癌死亡的主要原因之一。β-tubulin是抗微管药物的主要细胞靶点。已有的研究证明:βIII-tubulin高表达与非小细胞肺癌(non-small cell lung cancer, NSCLC)耐药有关。利用RNA干扰技术沉默耐紫杉醇A549细胞(A549/Taxol)中βIII-tubulin基因表达,探讨靶基因下调后对化疗药物紫杉醇的敏感性的变化以及细胞周期和细胞凋亡情况。方法 构建靶向βIII-tubulin的siRNA,以脂质体为载体介导βIII-tubulin siRNA转染A549/Taxol细胞,利用qRT-PCR检测细胞内βIII-tubulin mRNA的变化情况,并筛选出最佳干扰序列;Western blot法检测A549/Taxol细胞内βIII-tubulin蛋白表达的变化;MTT法检测转染后细胞株对紫杉醇敏感性的变化;流式细胞仪检测细胞周期和细胞凋亡的变化。结果 实时荧光qRT-PCR法显示转染后细胞株靶基因水平较对照组降低,其中βIII-tubulin siRNA-1序列抑制率最高为(87.73±4.87)%(P<0.01);Western blot显示转染后靶蛋白水平较对照组明显降低;MTT法表明紫杉醇处理转染后细胞株的细胞抑制率较对照组明显增加(51.77±4.60)%(P<0.01);细胞凋亡显示βIII-tubulin siRNA+Taxol组细胞早期凋亡率较对照组明显增加(P<0.01),两者的差异有统计学意义;细胞周期检测结果显示紫杉醇处理组的G2/M期细胞百分率高于对照组,且转染后紫杉醇处理组的细胞晚期凋亡率较对照组增加。结论 βIII-tubulin表达下调明显提高A549/Taxol细胞株对Taxol的敏感性。

  17. Down-regulation in human cancers of DRHC, a novel helicase-like gene from 17q25.1 that inhibits cell growth.

    Science.gov (United States)

    Nagai, H; Yabe, A; Mine, N; Mikami, I; Fujiwara, H; Terada, Y; Hirano, A; Tsuneizumi, M; Yokota, T; Emi, M

    2003-04-10

    Frequent observations of allelic loss in chromosomal band 17q25.1 in a variety of human cancers have suggested that one or more tumor suppressor genes are normally present in this region. Moreover, a locus responsible for hereditary focal non-epidermolytic palmoplantar keratoderma (tylosis oesophageal cancer; TOC), a condition associated with esophageal cancer, has been mapped to the same band. During efforts to sequence, by shot-gun methods, a 1 Mb target region that we had defined as the DNA segment harboring the putative tumor suppressor gene(s) involved in these events, we identified a novel cDNA, DRHC (down-regulated in human cancers), that showed reduced expression in 28 of 95 (29%) cell lines derived from a variety of human cancers. The full-length cDNA, 6275 bp long, was expressed predominantly in thymus and brain. The predicted 1942-amino-acid product exhibited significant sequence homology to yeast enzymes belonging to the DEAD-helicase superfamily, and appeared to be a Uvr/Rep helicase with a DEXDc consensus domain. Transfection of a DRHC expression vector inhibited growth of cancer cells in liquid medium or soft agar. The results suggest that loss of expression of DRHC may play a role in human carcinogenesis.

  18. Down-regulation of progesterone receptor membrane component 1 (PGRMC1 in peripheral nucleated blood cells associated with premature ovarian failure (POF and polycystic ovary syndrome (PCOS

    Directory of Open Access Journals (Sweden)

    Karlström Per-Olof

    2010-06-01

    Full Text Available Abstract Background Progesterone receptor membrane component 1 (PGRMC1 is a member of a progesterone-binding complex implicated in female reproduction. We aimed i to determine the natural expression of PGRMC1 in peripheral nucleated blood cells throughout the menstrual cycle and ii to investigate any association between PGRMC1 levels in leukocytes and conditions characterized by reduced fertility. Methods We analyzed PGRMC1 expression in peripheral leukocytes from 15 healthy cycling women over four weeks. Additionally, we determined PGRMC1 levels in samples from patients with premature ovarian failure (POF and polycystic ovary syndrome (PCOS as well as in healthy postmenopausal women and male controls. The levels of PGRMC1 protein in nucleated peripheral blood cells were quantified by Western blot analysis. Results PGRMC1 levels did not vary significantly throughout the menstrual cycle. We observed a significant down-regulation of PGRMC1 in postmenopausal women and in patients with premature ovarian failure (POF and polycystic ovary syndrome (PCOS when compared to early follicular phase of healthy women. Conclusion This study suggests that reduced levels of PGRMC1 in peripheral leukocytes are associated with perturbed ovulatory function.

  19. Nicotine-induced survival signaling in lung cancer cells is dependent on their p53 status while its down-regulation by curcumin is independent

    Directory of Open Access Journals (Sweden)

    Puliyappadamba Vineshkumar T

    2010-08-01

    Full Text Available Abstract Background Lung cancer is the most lethal cancer and almost 90% of lung cancer is due to cigarette smoking. Even though nicotine, one of the major ingredients of cigarette smoke and the causative agent for addiction, is not a carcinogen by itself, several investigators have shown that nicotine can induce cell proliferation and angiogenesis. We observed that the proliferative index of nicotine is different in the lung cancer cell lines H1299 (p53-/- and A549 (p53+/+ which indicates that the mode of up-regulation of survival signals by nicotine might be different in cells with and without p53. Results While low concentrations of nicotine induced activation of NF-κB, Akt, Bcl2, MAPKs, AP1 and IAPs in H1299, it failed to induce NF-κB in A549, and compared to H1299, almost 100 times higher concentration of nicotine was required to induce all other survival signals in A549. Transfection of WT-p53 and DN-p53 in H1299 and A549 respectively, reversed the mode of activation of survival signals. Curcumin down-regulated all the survival signals induced by nicotine in both the cells, irrespective of their p53 status. The hypothesis was confirmed when lower concentrations of nicotine induced NF-κB in two more lung cancer cells, Hop-92 and NCI-H522 with mutant p53 status. Silencing of p53 in A549 using siRNA made the cells susceptible to nicotine-induced NF-κB nuclear translocation as in A549 DN-p53 cells. Conclusions The present study reveals a detrimental role of nicotine especially in lung cancer patients with impaired p53 status and identifies curcumin as a potential chemopreventive.

  20. Interferon-inducible IFI16, a negative regulator of cell growth, down-regulates expression of human telomerase reverse transcriptase (hTERT gene.

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    Lynda Li Song

    Full Text Available BACKGROUND: Increased levels of interferon (IFN-inducible IFI16 protein (encoded by the IFI16 gene located at 1q22 in human normal prostate epithelial cells and diploid fibroblasts (HDFs are associated with the onset of cellular senescence. However, the molecular mechanisms by which the IFI16 protein contributes to cellular senescence-associated cell growth arrest remain to be elucidated. Here, we report that increased levels of IFI16 protein in normal HDFs and in HeLa cells negatively regulate the expression of human telomerase reverse transcriptase (hTERT gene. METHODOLOGY/PRINCIPAL FINDINGS: We optimized conditions for real-time PCR, immunoblotting, and telomere repeat amplification protocol (TRAP assays to detect relatively low levels of hTERT mRNA, protein, and telomerase activity that are found in HDFs. Using the optimized conditions, we report that treatment of HDFs with inhibitors of cell cycle progression, such as aphidicolin or CGK1026, which resulted in reduced steady-state levels of IFI16 mRNA and protein, was associated with increases in hTERT mRNA and protein levels and telomerase activity. In contrast, knockdown of IFI16 expression in cells increased the expression of c-Myc, a positive regulator of hTERT expression. Additionally, over-expression of IFI16 protein in cells inhibited the c-Myc-mediated stimulation of the activity of hTERT-luc-reporter and reduced the steady-state levels of c-Myc and hTERT. CONCLUSIONS/SIGNIFICANCE: These data demonstrated that increased levels of IFI16 protein in HDFs down-regulate the expression of hTERT gene. Our observations will serve basis to understand how increased cellular levels of the IFI16 protein may contribute to certain aging-dependent diseases.

  1. Interferon-Inducible IFI16, a Negative Regulator of Cell Growth, Down-Regulates Expression of Human Telomerase Reverse Transcriptase (hTERT) Gene

    Science.gov (United States)

    Shen, Hui; Duan, Xin; Alimirah, Fatouma; Choubey, Divaker

    2010-01-01

    Background Increased levels of interferon (IFN)-inducible IFI16 protein (encoded by the IFI16 gene located at 1q22) in human normal prostate epithelial cells and diploid fibroblasts (HDFs) are associated with the onset of cellular senescence. However, the molecular mechanisms by which the IFI16 protein contributes to cellular senescence-associated cell growth arrest remain to be elucidated. Here, we report that increased levels of IFI16 protein in normal HDFs and in HeLa cells negatively regulate the expression of human telomerase reverse transcriptase (hTERT) gene. Methodology/Principal Findings We optimized conditions for real-time PCR, immunoblotting, and telomere repeat amplification protocol (TRAP) assays to detect relatively low levels of hTERT mRNA, protein, and telomerase activity that are found in HDFs. Using the optimized conditions, we report that treatment of HDFs with inhibitors of cell cycle progression, such as aphidicolin or CGK1026, which resulted in reduced steady-state levels of IFI16 mRNA and protein, was associated with increases in hTERT mRNA and protein levels and telomerase activity. In contrast, knockdown of IFI16 expression in cells increased the expression of c-Myc, a positive regulator of hTERT expression. Additionally, over-expression of IFI16 protein in cells inhibited the c-Myc-mediated stimulation of the activity of hTERT-luc-reporter and reduced the steady-state levels of c-Myc and hTERT. Conclusions/Significance These data demonstrated that increased levels of IFI16 protein in HDFs down-regulate the expression of hTERT gene. Our observations will serve basis to understand how increased cellular levels of the IFI16 protein may contribute to certain aging-dependent diseases. PMID:20052289

  2. Down-regulation of Leucine-rich Repeats and Immunoglobulin-like Domain Proteins (LRIG1-3) in HP75 Pituitary Adenoma Cell Line

    Institute of Scientific and Technical Information of China (English)

    GUO Dongsheng; HAN Lin; SHU Kai; CHEN Jian; LEI Ting

    2007-01-01

    Three human leucine-rich repeats and immunoglobulin-like domains (LRIG) genes and proteins, named LRIG1-3, has been previously characterized and it was proposed that they may act as suppressors of tumor growth. The LRIG1 protein can inhibit the growth of tumors of glial cells and the down-regulation of the LRIG1 gene may be involved in the development and progression of the tumor. Real-time reverse transcription-polymerase chain reaction (RT-PCR) is a recently developed technique for quantitative assessment of specific RNA levels. In the current study, it was demonstrated that LRIG1-3 and EGFR mRNA was detected in human pituitary adenoma cell lines and a normal pituitary sample, with differences in the expression levels. Compared to the normal pituitary samples, the expression of LRIG1-3 in HP75 cell line was lower, but the expression of EGFR in HP75 cell line was higher. The results are consistent with LRIG1-3 being tumour suppressor genes, and LRIG genes decreasing the expression of EGFR. The ratio of EGFR/LRIG1 was increased at least 13-fold in HP75 cells compared with the normal pituitary cells, which was also the case for the ratio of EGFR/LRIG2 (14-fold increase in HP75) and EGFR/LRIG3 (11-fold increase in HP75). Further studies were needed to elucidate the explicit role of LRIG genes as negative regulators of oncogenesis in human pituitary adenoma.

  3. Down-regulation of G protein-coupled receptor 137 by RNA interference inhibits cell growth of two hepatoma cell lines.

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    Shao, Xin; Liu, Yong; Huang, Hai; Zhuang, Linyuan; Luo, Tianping; Huang, Huping; Ge, Xinguo

    2015-04-01

    G protein-coupled receptors (GPCRs) are important signal transduction mediators and pharmacological therapeutic targets. G protein-coupled receptor 137 (GPR137) was initially reported as a novel orphan GPCR around 10 years ago. Some orphan GPCRs have been implicated in cancer cell proliferation and migration. The aim of this study is to investigate the role of GPR137 in hepatocellular carcinoma (HCC). GPR137 is widely expressed in several human HCC cell lines, as determined by real-time PCR. We then applied lentivirus mediated RNA interference (RNAi) to knock down GPR137 expression in two HCC cell lines HepG2 and Bel7404. Depletion of GPR137 remarkably inhibited cell proliferation and colony formation capacity. Knockdown of GPR137 in HepG2 cells led to cell cycle arrest at G0/G1 phase and G2/M phase, and induced cell apoptosis, as determined by flow cytometry analysis, which contributed to cell growth inhibition. Our findings suggested that GPR137 could facilitate HCC cell proliferation and thus promote hepatocarcinogenesis.

  4. Reduced expression of Slit2 in renal cell carcinoma.

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    Ma, Wei-Jie; Zhou, Yu; Lu, Dan; Dong, Dong; Tian, Xiao-Jun; Wen, Jie-Xi; Zhang, Jun

    2014-01-01

    Slit2, initially identified as an important axon guidance molecule in the nervous system, was suggested to be involved in multiple cellular processes. Recently, Slit2 was reported to function as a potential tumor suppressor in diverse tumors. In this study, we systematically analyzed the expression level of Slit2 in renal cell carcinoma. Compared to paired adjacent non-malignant tissues, both Slit2 mRNA and protein expression were significantly down-regulated in renal cell carcinoma (RCC). Methylation-specific PCR showed that Slit2 promoter was methylated in two renal carcinoma cell lines. Pharmacologic demethylation dramatically induced Slit2 expression in cancer cell lines with weak expression of Slit2. Besides, bisulfite genomic sequencing confirmed that dense methylation existed in Slit2 promoter. Furthermore, in paired RCC samples, Slit2 methylation was observed in 8 out of 38 patients (21.1 %), which was well correlated with the down-regulation of Slit2 in RCC. Therefore, Slit2 may also be a potential tumor suppressor in RCC, which is down-regulated in RCC partially due to promoter methylation.

  5. Suppressive Effects of Plumbagin on Invasion and Migration of Breast Cancer Cells via the Inhibition of STAT3 Signaling and Down-regulation of Inflammatory Cytokine Expressions

    Institute of Scientific and Technical Information of China (English)

    Wei Yan; Bing Tu; Yun-yun Liu; Ting-yu Wang; Han Qiao; Zan-jing Zhai; Hao-wei Li; Ting-ting Tang

    2013-01-01

    Objective:The aim of this study was to investigate the effects of plumbagin (PL), a naphthoquinone derived from the medicinal plant plumbago zeylanica, on the invasion and migration of human breast cancer cells. Methods:Human breast cancer MDA-MB-231SArfp cells were treated with different concentrations of plum-bagin for 24 h. The effects of plumbagin on the migration and invasion were observed by a transwell method. The expressions of IL-1α, IL-1β, IL-6, IL-8, TGF-β, TNFα, MMP-2 and MMP-9 mRNA in M DA-MB-231SArfp cells were detected using Real-Time PCR. MDA-MB-231SArfp cells were treated with plumbagin at different concentrations for 45 minutes. The activation of STAT3 was detected by western blot. Following this analysis, STAT3 in MDA-MB-231SArfp cells was knocked out using specific siRNA. mRNA levels of IL-1α, TGF-β, MMP-2 and MMP-9 were then detected. Consequently, MDA-MB-231SArfp cells were injected intracardially into BALB/c nude mice to construct a breast cancer bone metastatic model. The mice were injected intra-peritoneally with plumbagin. Non-invasive in vivo monitoring, X-ray imaging and histological staining were performed to investigate the effects of plumbagin on the invasion and migration of breast cancer cells in vivo. Results: The in vitro results showed that plumbagin could suppress the migration and invasion of breast cancer cells and down-regulate mRNA expressions of IL-1α, TGF-β, MMP-2 and MMP-9. Western blotting demonstrated that plumbagin inhibited the activation of STAT3 signaling in MDA-MB-231SArfp cells. The inactivation of STAT3 was found to have an inhibitory effect on the expressions of IL-1α, TGF-β, MMP-2 and MMP-9. In vivo studies showed that plumbagin inhibited the metastasis of breast cancer cells and decreased osteolytic bone metastases, as well as the secretion of MMP-2 and MMP-9 by tumor cells at metastatic lesions. Conclusions:Plumbagin can suppress the invasion and migration of breast cancer cells via the inhibition

  6. Minocycline enhances mitomycin C-induced cytotoxicity through down-regulating ERK1/2-mediated Rad51 expression in human non-small cell lung cancer cells.

    Science.gov (United States)

    Ko, Jen-Chung; Wang, Tai-Jing; Chang, Po-Yuan; Syu, Jhan-Jhang; Chen, Jyh-Cheng; Chen, Chien-Yu; Jian, Yun-Ting; Jian, Yi-Jun; Zheng, Hao-Yu; Chen, Wen-Ching; Lin, Yun-Wei

    2015-10-01

    Minocycline is a semisynthetic tetracycline derivative; it has anti-inflammatory and anti-cancer effects distinct from its antimicrobial function. However, the molecular mechanism of minocycline-induced cytotoxicity in non-small cell lung cancer (NSCLC) cells has not been identified. Rad51 plays a central role in homologous recombination and high levels of Rad51 expression are observed in chemo- or radioresistant carcinomas. Our previous studies have shown that the MKK1/2-ERK1/2 signal pathway maintains the expression of Rad51 in NSCLC cells. In this study, minocycline treatment inhibited cell viability and proliferation of two NSCLC cells, A549 and H1975. Treatment with minocycline decreased Rad51 mRNA and protein levels through MKK1/2-ERK1/2 inactivation. Furthermore, expression of constitutively active MKK1 (MKK1-CA) vectors significantly rescued the decreased Rad51 protein and mRNA levels in minocycline-treated NSCLC cells. However, combined treatment with MKK1/2 inhibitor U0126 and minocycline further decreased the Rad51 expression and cell viability of NSCLC cells. Knocking down Rad51 expression by transfection with small interfering RNA of Rad51 enhanced the cytotoxicity and cell growth inhibition of minocycline. Mitomycin C (MMC) is typically used as a first or second line regimen to treat NSCLC. Compared to a single agent alone, MMC combined with minocycline resulted in cytotoxicity and cell growth inhibition synergistically in NSCLC cells, accompanied with reduced activation of phospho-ERK1/2, and reduced Rad51 protein levels. Overexpression of MKK1-CA or Flag-tagged Rad51 could reverse the minocycline and MMC-induced synergistic cytotoxicity. These findings may have implications for the rational design of future drug regimens incorporating minocycline and MMC for the treatment of NSCLC.

  7. Astaxanthin down-regulates Rad51 expression via inactivation of AKT kinase to enhance mitomycin C-induced cytotoxicity in human non-small cell lung cancer cells.

    Science.gov (United States)

    Ko, Jen-Chung; Chen, Jyh-Cheng; Wang, Tai-Jing; Zheng, Hao-Yu; Chen, Wen-Ching; Chang, Po-Yuan; Lin, Yun-Wei

    2016-04-01

    Astaxanthin has been demonstrated to exhibit a wide range of beneficial effects, including anti-inflammatory and anti-cancer properties. However, the molecular mechanism of astaxanthin-induced cytotoxicity in non-small cell lung cancer (NSCLC) cells has not been identified. Rad51 plays a central role in homologous recombination, and studies show that chemo-resistant carcinomas exhibit high levels of Rad51 expression. In this study, astaxanthin treatment inhibited cell viability and proliferation of two NSCLC cells, A549 and H1703. Astaxanthin treatment (2.5-20 μM) decreased Rad51 expression and phospho-AKT(Ser473) protein level in a time and dose-dependent manner. Furthermore, expression of constitutively active AKT (AKT-CA) vector rescued the decreased Rad51 mRNA and protein levels in astaxanthin-treated NSCLC cells. Combined treatment with phosphatidylinositol 3-kinase (PI3K) inhibitors (LY294002 or wortmannin) further decreased the Rad51 expression in astaxanthin-exposed A549 and H1703 cells. Knockdown of Rad51 expression by transfection with si-Rad51 RNA or cotreatment with LY294002 further enhanced the cytotoxicity and cell growth inhibition of astaxanthin. Additionally, mitomycin C (MMC) as an anti-tumor antibiotic is widely used in clinical NSCLC chemotherapy. Combination of MMC and astaxanthin synergistically resulted in cytotoxicity and cell growth inhibition in NSCLC cells, accompanied with reduced phospho-AKT(Ser473) level and Rad51 expression. Overexpression of AKT-CA or Flag-tagged Rad51 reversed the astaxanthin and MMC-induced synergistic cytotoxicity. In contrast, pretreatment with LY294002 further decreased the cell viability in astaxanthin and MMC co-treated cells. In conclusion, astaxanthin enhances MMC-induced cytotoxicity by decreasing Rad51 expression and AKT activation. These findings may provide rationale to combine astaxanthin with MMC for the treatment of NSCLC.

  8. MicroRNA-16 is down-regulated in mutated FLT3 expressing murine myeloid FDC-P1 cells and interacts with Pim-1.

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    Kyu-Tae Kim

    Full Text Available Activating mutations in the receptor tyrosine kinase FLT3 are one of the most frequent somatic mutations in acute myeloid leukemia (AML. Internal tandem duplications of the juxtamembrane region of FLT3 (FLT3/ITD constitutively activate survival and proliferation pathways, and are associated with a poor prognosis in AML. We suspected that alteration of small non-coding microRNA (miRNA expression in these leukemia cells is involved in the transformation process and used miRNA microarrays to determine the miRNA signature from total RNA harvested from FLT3/ITD expressing FDC-P1 cells (FD-FLT3/ITD. This revealed that a limited set of miRNAs appeared to be affected by expression of FLT3/ITD compared to the control group consisting of FDC-P1 parental cells transfected with an empty vector (FD-EV. Among differentially expressed miRNAs, we selected miR-16, miR-21 and miR-223 to validate the microarray data by quantitative real-time RT-PCR showing a high degree of correlation. We further analyzed miR-16 expression with FLT3 inhibitors in FLT3/ITD expressing cells. MiR-16 was found to be one of most significantly down-regulated miRNAs in FLT3/ITD expressing cells and was up-regulated upon FLT3 inhibition. The data suggests that miR-16 is acting as a tumour suppressor gene in FLT3/ITD-mediated leukemic transformation. Whilst miR-16 has been reported to target multiple mRNAs, computer models from public bioinformatic resources predicted a potential regulatory mechanism between miR-16 and Pim-1 mRNA. In support of this interaction, miR-16 was shown to suppress Pim-1 reporter gene expression. Further, our data demonstrated that over-expression of miR-16 mimics suppressed Pim-1 expression in FD-FLT3/ITD cells suggesting that increased miR-16 expression contributes to depletion of Pim-1 after FLT3 inhibition and that miR-16 repression may be associated with up-regulated Pim-1 in FLT3/ITD expressing cells.

  9. Genes encoding Pir51,Beclin 1,RbAp48 and aldolase b are up or down-regulated in human primary hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Hai Song; Shuang-Luo Xia; Cheng Liao; Yi-Liang Li; Yi-Fei Wang; Tsai-Ping Li; Mu-Jun Zhao

    2004-01-01

    AIM: To reveal new tumor markers and target genes from differentially expressed genes of primary tumor samples using cDNA microarray.METHODS: The 33p labeled cDNAs were synthesized by reverse transcription of message RNA from the liver cancerous tissue and adjacent non-cancerous liver tissue from the same patient and used to hybridize to LifeGrid 1.0cDNA microarray blot containing 8400 known and unique human cDNA gene targets, and an expression profile of genes was produced in one paired human liver tumor tissue.After a global analysis of gene expression of 8400 genes,we selected some genes to confirm the differential expression using Northern blot and RT-PCR.RESULTS: Parallel analysis of the hybridized signals enabled us to get an expression profile of genes in which about 500genes were differentially expressed in the paired liver tumor tissues. We identified 4 genes, the expression of three(Beclin 1, RbAp48 and Pir51) were increased and one (aldolase b)was decreased in liver tumor tissues. In addition, the expression of these genes in 6 hepatoma cell lines was also showed by RT-PCR analysis.CONCLUSION: cDNA microarray permits a high throughput identification of changes in gene expression. The genes encoding Beclin 1, RbAp48, Pir51 and aldolase b are first reported that may be related with hepatocarcinoma.

  10. The expression of VEGF-A is down regulated in peripheral blood mononuclear cells of patients with secondary progressive multiple sclerosis.

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    Ellen Iacobaeus

    Full Text Available BACKGROUND: Most patients with relapsing-remitting multiple sclerosis (RRMS eventually enter a secondary progressive (SPMS phase, characterized by increasing neurological disability. The mechanisms underlying transition to SPMS are unknown and effective treatments and biomarkers are lacking. Vascular endothelial growth factor-A (VEGF-A is an angiogenic factor with neuroprotective effects that has been associated with neurodegenerative diseases. SPMS has a prominent neurodegenerative facet and we investigated a possible role for VEGF-A during transition from RRMS to SPMS. METHODOLOGY/PRINCIPAL FINDINGS: VEGF-A mRNA expression in peripheral blood mononuclear (PBMC and cerebrospinal fluid (CSF cells from RRMS (n = 128, SPMS (n = 55 and controls (n = 116 were analyzed using real time PCR. We demonstrate reduced expression of VEGF-A mRNA in MS CSF cells compared to controls (p<0.001 irrespective of disease course and expression levels are restored by natalizumab treatment(p<0.001. VEGF-A was primarily expressed in monocytes and our CSF findings in part may be explained by effects on relative monocyte proportions. However, VEGF-A mRNA expression was also down regulated in the peripheral compartment of SPMS (p<0.001, despite unchanged monocyte counts, demonstrating a particular phenotype differentiating SPMS from RRMS and controls. A possible association of allelic variability in the VEGF-A gene to risk of MS was also studied by genotyping for six single nucleotide polymorphisms (SNPs in MS (n = 1114 and controls (n = 1234, which, however, did not demonstrate any significant association between VEGF-A alleles and risk of MS. CONCLUSIONS/SIGNIFICANCE: Expression of VEGF-A in CSF cells is reduced in MS patients compared to controls irrespective of disease course. In addition, SPMS patients display reduced VEGF-A mRNA expression in PBMC, which distinguish them from RRMS and controls. This indicates a possible role for VEGF-A in the mechanisms regulating

  11. Down-regulation of 5-HT1B and 5-HT1D receptors inhibits proliferation, clonogenicity and invasion of human pancreatic cancer cells.

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    Nilgun Gurbuz

    Full Text Available Pancreatic ductal adenocarcinoma is characterized by extensive local tumor invasion, metastasis and early systemic dissemination. The vast majority of pancreatic cancer (PaCa patients already have metastatic complications at the time of diagnosis, and the death rate of this lethal type of cancer has increased over the past decades. Thus, efforts at identifying novel molecularly targeted therapies are priorities. Recent studies have suggested that serotonin (5-HT contributes to the tumor growth in a variety of cancers including prostate, colon, bladder and liver cancer. However, there is lack of evidence about the impact of 5-HT receptors on promoting pancreatic cancer. Having considered the role of 5-HT-1 receptors, especially 5-HT1B and 5-HT1D subtypes in different types of malignancies, the aim of this study was to investigate the role of 5-HT1B and 5-HT1D receptors in PaCa growth and progression and analyze their potential as cytotoxic targets. We found that knockdown of 5-HT1B and 5-HT1D receptors expression, using specific small interfering RNA (siRNA, induced significant inhibition of proliferation and clonogenicity of PaCa cells. Also, it significantly suppressed PaCa cells invasion and reduced the activity of uPAR/MMP-2 signaling and Integrin/Src/Fak-mediated signaling, as integral tumor cell pathways associated with invasion, migration, adhesion, and proliferation. Moreover, targeting 5-HT1B and 5-HT1D receptors down-regulates zinc finger ZEB1 and Snail proteins, the hallmarks transcription factors regulating epithelial-mesenchymal transition (EMT, concomitantly with up-regulating of claudin-1 and E-Cadherin. In conclusion, our data suggests that 5-HT1B- and 5-HT1D-mediated signaling play an important role in the regulation of the proliferative and invasive phenotype of PaCa. It also highlights the therapeutic potential of targeting of 5-HT1B/1D receptors in the treatment of PaCa, and opens a new avenue for biomarkers identification

  12. Quinacrine induces apoptosis in human leukemia K562 cells via p38 MAPK-elicited BCL2 down-regulation and suppression of ERK/c-Jun-mediated BCL2L1 expression

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    Changchien, Jung-Jung; Chen, Ying-Jung; Huang, Chia-Hui [Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung 804, Taiwan (China); Cheng, Tian-Lu [Department of Biomedical Science and Environmental Biology, Kaohsiung Medical University, Kaohsiung 807, Taiwan (China); Lin, Shinne-Ren [Department of Medicinal and Applied Chemistry, Kaohsiung Medical University, Kaohsiung 807, Taiwan (China); Chang, Long-Sen, E-mail: lschang@mail.nsysu.edu.tw [Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung 804, Taiwan (China); Department of Biotechnology, Kaohsiung Medical University, Kaohsiung 807, Taiwan (China)

    2015-04-01

    Although previous studies have revealed the anti-cancer activity of quinacrine, its effect on leukemia is not clearly resolved. We sought to explore the cytotoxic effect and mechanism of quinacrine action in human leukemia K562 cells. Quinacrine induced K562 cell apoptosis accompanied with ROS generation, mitochondrial depolarization, and down-regulation of BCL2L1 and BCL2. Upon exposure to quinacrine, ROS-mediated p38 MAPK activation and ERK inactivation were observed in K562 cells. Quinacrine-induced cell death and mitochondrial depolarization were suppressed by the p38MAPK inhibitor SB202190 and constitutively active MEK1 over-expression. Activation of p38 MAPK was shown to promote BCL2 degradation. Further, ERK inactivation suppressed c-Jun-mediated transcriptional expression of BCL2L1. Over-expression of BCL2L1 and BCL2 attenuated quinacrine-evoked mitochondrial depolarization and rescued the viability of quinacrine-treated cells. Taken together, our data indicate that quinacrine-induced K562 cell apoptosis is mediated through mitochondrial alterations triggered by p38 MAPK-mediated BCL2 down-regulation and suppression of ERK/c-Jun-mediated BCL2L1 expression. - Highlights: • Quinacrine induces K562 cell apoptosis via down-regulation of BCL2 and BCL2L1. • Quinacrine induces p38 MAPK activation and ERK inactivation in K562 cells. • Quinacrine elicits p38 MAPK-mediated BCL2 down-regulation. • Quinacrine suppresses ERK/c-Jun-mediated BCL2L1 expression.

  13. MicroRNA-Mediated Down-Regulation of Apoptosis Signal-Regulating Kinase 1 (ASK1 Attenuates the Apoptosis of Human Mesenchymal Stem Cells (MSCs Transplanted into Infarcted Heart

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    Chang Youn Lee

    2016-10-01

    Full Text Available Stem cell therapy using adult stem cells, such as mesenchymal stem cells (MSCs has produced some promising results in treating the damaged heart. However, the low survival rate of MSCs after transplantation is still one of the crucial factors that limit the therapeutic effect of stem cells. In the damaged heart, oxidative stress due to reactive oxygen species (ROS production can cause the death of transplanted MSCs. Apoptosis signal-regulating kinase 1 (ASK1 has been implicated in the development of oxidative stress-related pathologic conditions. Thus, we hypothesized that down-regulation of ASK1 in human MSCs (hMSCs might attenuate the post-transplantation death of MSCs. To test this hypothesis, we screened microRNAs (miRNAs based on a miRNA-target prediction database and empirical data and investigated the anti-apoptotic effect of selected miRNAs on human adipose-derived stem cells (hASCs and on rat myocardial infarction (MI models. Our data indicated that miRNA-301a most significantly suppressed ASK1 expression in hASCs. Apoptosis-related genes were significantly down-regulated in miRNA-301a-enriched hASCs exposed to hypoxic conditions. Taken together, these data show that miRNA-mediated down-regulation of ASK1 protects MSCs during post-transplantation, leading to an increase in the efficacy of MSC-based cell therapy.

  14. Advanced glycation end products promote human aortic smooth muscle cell calcification in vitro via activating NF-κB and down-regulating IGF1R expression

    Institute of Scientific and Technical Information of China (English)

    Yi WANG; Zhen-yu ZHANG; Xiao-qing CHEN; Xiang WANG; Heng CAO; Shao-wen LIU

    2013-01-01

    Aim:To investigate the effects of advanced glycation end products (AGEs) on calcification in human aortic smooth muscle cells (HASMCs) in vitro and the underlying mechanisms.Methods:AGEs were artificially prepared.Calcification of HASMCs was induced by adding inorganic phosphate (Pi,2 mmol/L) in the media,and observed with Alizarin red staining.The calcium content in the supernatant was measured using QuantiChrome Calcium Assay Kit.Expression of the related mRNAs and proteins was analyzed using real-time PCR and Western blot,respectively.Chromatin immunoprecipitation (ChIP) assay was used to detect the binding of NF-κB to the putative IGF1R promoter.Results:AGEs (100 μg/mL) significantly enhanced Pi-induced calcification and the levels of osteocalcin and Cbfα1 in HASMCs.Furthermore,the treatment decreased the expression of insulin-like growth factor 1 receptor (IGF1R).Over-expression of IGF1R in HASMCs suppressed the AGEs-induced increase in calcium deposition.When IGF1R expression was knocked down in HASMCs,AGEs did not enhance the calcium deposition.Meanwhile,AGEs time-dependently decreased the amounts of IκBα and Flag-tagged p65 in the cytoplasmic extracts,and increased the amount of nuclear p65 in HASMCs.In the presence of NF-κB inhibitor PDTC (50 μmol/L),the AGEs-induced increase in calcium deposition was blocked.Over-expression of p65 significantly enhanced Pi-induced mineralization,but suppressed IGF1R mRNA level.Knockdown of p65 suppressed the AGEs-induced increase in calcium deposition,and rescued the IGF1R expression.The ChIP analysis revealed that NF-κB bound the putative IGF1R promoter at position-230 to-219 bp.The inhibition of IGF1R by NF-κB was abolished when IGF1R reporter plasmid contained mutated binding sequence for NF-κB or an NF-κB reporter vector.Conclusion:The results demonstrate that AGEs promote calcification of human aortic smooth muscle cells in vitro via activation of NF-κB and down-regulation of IGF1R expression.

  15. Hypoxia in Tumor Angiogenesis and Metastasis: Evaluation of VEGF and MMP Over-expression and Down-Regulation of HIF-1alpha with RNAi in Hypoxic Tumor Cells

    Science.gov (United States)

    Shah, Shruti

    Background: As tumor mass grows beyond a few millimeters in diameter, the angiogenic "switch" is turned on leading to recruitment of blood vessels from surrounding artery and veins. However, the tumor mass is poorly perfused and there are pockets of hypoxia or lower oxygen concentrations relative to normal tissue. Hypoxia-inducing factor-1a (HIF-1a), a transcription factor, is activated when the oxygen concentration is low. Upon activation of HIF-1a, a number of other genes also turn on that allows the tumor to become more aggressive and resistant to therapy. Purpose: The main objectives of this study were to evaluate the effect of hypoxia-induced HIF-1a followed by over-expression of angiogenic and metastatic markers in tumor cells and down-regulation of HIF-1a using nanoparticle-delivered RNA interference therapy. Methods: Human ovarian (SKOV3) and breast (MDA-MB-231) adenocarcinoma cells were incubated under normoxic and hypoxic conditions. Following hypoxia treatment of the cells, HIF-1α, vascular endothelial growth factor (VEGF), matrix metalloproteinase 2 (MMP-2), and MMP-9 expression was analyzed qualitatively and quantitatively. For intracellular delivery of HIF-1a gene silencing small interfering RNA (siRNA), type B gelatin nanoparticles were fabricated using the solvent displacement method and the surface was modified with poly(ethylene glycol) (PEG, Mol. wt. 2kDa). Cellular uptake and distribution of the nanoparticles was observed with Cy3-siRNA loaded, FITC-conjugated gelatin nanoparticles. Cytotoxicity of the nanoparticle formulations was evaluated in both the cell lines. siRNA was transfected in the gelatin nanoparticles under hypoxic conditions. Total cellular protein and RNA were extracted for analysis of HIF1a, VEGF, MMP-2 and MMP-9 expression. Results: MDA-MB-231 and SKOV3 cells show increased expression of HIF1a under hypoxic conditions compared to baseline levels at normoxic conditions. ELISA and western blots of VEGF, MMP-2 and MMP-9 appear to

  16. Giant basal cell carcinoma Carcinoma basocelular gigante

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    Nilton Nasser

    2012-06-01

    Full Text Available The basal cell carcinoma is the most common skin cancer but the giant vegetating basal cell carcinoma reaches less than 0.5 % of all basal cell carcinoma types. The Giant BCC, defined as a lesion with more than 5 cm at its largest diameter, is a rare form of BCC and commonly occurs on the trunk. This patient, male, 42 years old presents a Giant Basal Cell Carcinoma which reaches 180 cm2 on the right shoulder and was negligent in looking for treatment. Surgical treatment was performed and no signs of dissemination or local recurrence have been detected after follow up of five years.O carcinoma basocelular é o tipo mais comum de câncer de pele, mas o carcinoma basocelular gigante vegetante não atinge 0,5% de todos os tipos de carcinomas basocelulares. O Carcinoma Basocelular Gigante, definido como lesão maior que 5 cm no maior diâmetro, é uma forma rara de carcinoma basocelular e comumente ocorre no tronco. Este paciente apresenta um Carcinoma Basocelular Gigante com 180cm² no ombro direito e foi negligente em procurar tratamento. Foi realizado tratamento cirúrgico e nenhum sinal de disseminação ou recorrência local foi detectada após 5 anos.

  17. 5-Bromo-3-(3-hydroxyprop-1-ynyl-2H-pyran-2-one targets prostate cancer cells by down-regulating inflammation-related genes

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    Zhao-Yang Wang

    2016-06-01

    Full Text Available The present study was performed to examine the effect of 5-bromo-3-(3-hydroxyprop-1-ynyl-2H-pyran-2-one (BHP on the rate of cell proliferation and apoptosis induction in the PC3, human prostate carcinoma cell line. The cell viability was assayed by using sulphorhodamine B staining and apoptosis by annexin V and flow cytometry analyses. The results revealed that BHP treatment in PC3 cells caused a significant reduction in the rate of cell proliferation in dose- and time-dependent manner. Compared to the un-treated cells, the formation of HUVEC tubes was markedly inhibited on treatment with BHP at a concentration of 30 µM. Further investigation revealed the expression of HMGB1, IL-6 and IL-8, pro-inflammatory cytokines was also inhibited on treatment with BHP. Therefore, BHP treatment plays an important role in inducing apoptosis in the prostrate cells and can be of therapeutic value for the prostate cancer treatment.

  18. Down-regulation of mitogen-activated protein kinases and nuclear factor-κB signaling is involved in rapamycin suppression of TLR2-induced inflammatory response in monocytic THP-1 cells.

    Science.gov (United States)

    Sun, Ruili; Zhang, Yi; Ma, Shijiang; Qi, Hengtian; Wang, Mingyong; Duan, Juhong; Ma, Shujun; Zhu, Xiaofei; Li, Guancheng; Wang, Hui

    2015-10-01

    Tripalmitoyl-S-glycero-Cys-(Lys) 4 (Pam3CSK4) interacted with TLR2 induces inflammatory responses through the mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NF-κB) signal pathway. Rapamycin can suppress TLR-induced inflammatory responses; however, the detailed molecular mechanism is not fully understood. Here, the mechanism by which rapamycin suppresses TLR2-induced inflammatory responses was investigated. It was found that Pam3CSK4-induced pro-inflammatory cytokines were significantly down-regulated at both the mRNA and protein levels in THP-1 cells pre-treated with various concentrations of rapamycin. Inhibition of phosphatidylinositol 3-kinase/protein kinase-B (PI3K/AKT) signaling did not suppress the expression of pro-inflammatory cytokines, indicating that the immunosuppression mediated by rapamycin in THP1 cells is independent of the PI3K/AKT pathway. RT-PCR showed that Erk and NF-κB signal pathways are related to the production of pro-inflammatory cytokines. Inhibition of Erk or NF-κB signaling significantly down-regulated production of pro-inflammatory cytokines. Additionally, western blot showed that pre-treatment of THP-1 cells with rapamycin down-regulates MAPKs and NF-κB signaling induced by Pam3CSK4 stimulation, suggesting that rapamycin suppresses Pam3CSK4-induced pro-inflammatory cytokines via inhibition of TLR2 signaling. It was concluded that rapamycin suppresses TLR2-induced inflammatory responses by down-regulation of Erk and NF-κB signaling.

  19. Inhibition of H3K9me2 Reduces Hair Cell Regeneration after Hair Cell Loss in the Zebrafish Lateral Line by Down-Regulating the Wnt and Fgf Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Dongmei eTang

    2016-05-01

    Full Text Available The activation of neuromast supporting cell (SC proliferation leads to hair cell (HC regeneration in the zebrafish lateral line. Epigenetic mechanisms have been reported that regulate HC regeneration in the zebrafish lateral line, but the role of H3K9me2 in HC regeneration after HC loss remains poorly understood. In this study, we focused on the role of H3K9me2 in HC regeneration following neomycin-induced HC loss. To investigate the effects of H3K9me2 in HC regeneration, we took advantage of the G9a/GLP-specific inhibitor BIX01294 that significantly reduces the dimethylation of H3K9. We found that BIX01294 significantly reduced HC regeneration after neomycin-induced HC loss in the zebrafish lateral line. BIX01294 also significantly reduced the proliferation of neuromast cells and led to fewer SCs in the lateral line. In situ hybridization showed that BIX01294 significantly down-regulated the Wnt and Fgf signaling pathways, which resulted in reduced SC proliferation and HC regeneration in the neuromasts of the lateral line. Altogether, our results suggest that down-regulation of H3K9me2 significantly decreases HC regeneration after neomycin-induced HC loss through inactivation of the Wnt/β-catenin and Fgf signaling pathways. Thus H3K9me2 plays a critical role in HC regeneration.

  20. Inhibition of H3K9me2 Reduces Hair Cell Regeneration after Hair Cell Loss in the Zebrafish Lateral Line by Down-Regulating the Wnt and Fgf Signaling Pathways

    Science.gov (United States)

    Tang, Dongmei; Lin, Qin; He, Yingzi; Chai, Renjie; Li, Huawei

    2016-01-01

    The activation of neuromast (NM) supporting cell (SC) proliferation leads to hair cell (HC) regeneration in the zebrafish lateral line. Epigenetic mechanisms have been reported that regulate HC regeneration in the zebrafish lateral line, but the role of H3K9me2 in HC regeneration after HC loss remains poorly understood. In this study, we focused on the role of H3K9me2 in HC regeneration following neomycin-induced HC loss. To investigate the effects of H3K9me2 in HC regeneration, we took advantage of the G9a/GLP-specific inhibitor BIX01294 that significantly reduces the dimethylation of H3K9. We found that BIX01294 significantly reduced HC regeneration after neomycin-induced HC loss in the zebrafish lateral line. BIX01294 also significantly reduced the proliferation of NM cells and led to fewer SCs in the lateral line. In situ hybridization showed that BIX01294 significantly down-regulated the Wnt and Fgf signaling pathways, which resulted in reduced SC proliferation and HC regeneration in the NMs of the lateral line. Altogether, our results suggest that down-regulation of H3K9me2 significantly decreases HC regeneration after neomycin-induced HC loss through inactivation of the Wnt/β-catenin and Fgf signaling pathways. Thus H3K9me2 plays a critical role in HC regeneration. PMID:27303264

  1. Down-regulation of PERK enhances resistance to ionizing radiation

    Energy Technology Data Exchange (ETDEWEB)

    Oommen, Deepu, E-mail: oommen1978@gmail.com; Prise, Kevin M.

    2013-11-08

    Highlights: •PERK enhances the sensitivity of cancer cells to ionizing radiation. •Down-regulation of PERK results in enhanced DNA repair. •Ionizing radiation-induced apoptosis is inhibited in PERK-down regulated cancer cells. -- Abstract: Although, ionizing radiation (IR) has been implicated to cause stress in endoplasmic reticulum (ER), how ER stress signaling and major ER stress sensors modulate cellular response to IR is unclear. Protein kinase RNA-like endoplasmic reticulum kinase (PERK) is an ER transmembrane protein which initiates unfolded protein response (UPR) or ER stress signaling when ER homeostasis is disturbed. Here, we report that down-regulation of PERK resulted in increased clonogenic survival, enhanced DNA repair and reduced apoptosis in irradiated cancer cells. Our study demonstrated that PERK has a role in sensitizing cancer cells to IR.

  2. A novel neuron-enriched protein SDIM1 is down regulated in Alzheimer's brains and attenuates cell death induced by DNAJB4 over-expression in neuro-progenitor cells

    Directory of Open Access Journals (Sweden)

    Lei Joy X

    2011-01-01

    Full Text Available Abstract Background Molecular changes in multiple biological processes contribute to the development of chronic neurodegeneration such as late onset Alzheimer's disease (LOAD. To discover how these changes are reflected at the level of gene expression, we used a subtractive transcription-based amplification of mRNA procedure to identify novel genes that have altered expression levels in the brains of Alzheimer's disease (AD patients. Among the genes altered in expression level in AD brains was a transcript encoding a novel protein, SDIM1, that contains 146 amino acids, including a typical signal peptide and two transmembrane domains. Here we examined its biochemical properties and putative roles in neuroprotection/neurodegeneration. Results QRT-PCR analysis of additional AD and control post-mortem human brains showed that the SDIM1 transcript was indeed significantly down regulated in all AD brains. SDIM1 is more abundant in NT2 neurons than astrocytes and present throughout the cytoplasm and neural processes, but not in the nuclei. In NT2 neurons, it is highly responsive to stress conditions mimicking insults that may cause neurodegeneration in AD brains. For example, SDIM1 was significantly down regulated 2 h after oxygen-glucose deprivation (OGD, though had recovered 16 h later, and also appeared significantly up regulated compared to untreated NT2 neurons. Overexpression of SDIM1 in neuro-progenitor cells improved cells' ability to survive after injurious insults and its downregulation accelerated cell death induced by OGD. Yeast two-hybrid screening and co-immunoprecipitation approaches revealed, both in vitro and in vivo, an interaction between SDIM1 and DNAJB4, a heat shock protein hsp40 homolog, recently known as an enhancer of apoptosis that also interacts with the mu opioid receptor in human brain. Overexpression of DNAJB4 alone significantly reduced cell viability and SDIM1 co-overexpression was capable of attenuating the cell death

  3. TRAIL sensitize MDR cells to MDR-related drugs by down-regulation of P-glycoprotein through inhibition of DNA-PKcs/Akt/GSK-3β pathway and activation of caspases

    Directory of Open Access Journals (Sweden)

    Kim Dong-Wan

    2010-07-01

    Full Text Available Abstract Background The development of new modulator possessing high efficacy, low toxicity and high selectivity is a pivotal approach to overcome P-glycoprotein (P-gp mediated multidrug resistance (MDR in cancer treatment. In this study, we suggest a new molecular mechanism that TRAIL (tumor necrosis factor-related apoptosis-inducing ligand down-regulates P-glycoprotein (P-gp through inhibition of DNA-PKcs/Akt/GSK-3β pathway and activation of caspases and thereby sensitize MDR cells to MDR-related drugs. Results MDR variants, CEM/VLB10-2, CEM/VLB55-8 and CEM/VLB100 cells, with gradually increased levels of P-gp derived from human lymphoblastic leukemia CEM cells, were gradually more susceptible to TRAIL-induced apoptosis and cytotoxicity than parental CEM cells. The P-gp level of MDR variants was positively correlated with the levels of DNA-PKcs, pAkt, pGSK-3β and c-Myc as well as DR5 and negatively correlated with the level of c-FLIPs. Hypersensitivity of CEM/VLB100 cells to TRAIL was accompanied by the activation of mitochondrial apoptotic pathway as well as the activation of initiator caspases. In addition, TRAIL-induced down-regulation of DNA-PKcs/Akt/GSK-3β pathway and c-FLIP and up-regulation of cell surface expression of death receptors were associated with the increased susceptibility to TRAIL of MDR cells. Moreover, TRAIL inhibited P-gp efflux function via caspase-3-dependent degradation of P-gp as well as DNA-PKcs and subsequently sensitized MDR cells to MDR-related drugs such as vinblastine and doxorubicin. We also found that suppression of DNA-PKcs by siRNA enhanced the susceptibility of MDR cells to vincristine as well as TRAIL via down-regulation of c-FLIP and P-gp expression and up-regulation of DR5. Conclusion This study showed for the first time that the MDR variant of CEM cells was hypersensitive to TRAIL due to up-regulation of DR5 and concomitant down-regulation of c-FLIP, and degradation of P-gp and DNA-PKcs by

  4. Ghost cell odontogenic carcinoma.

    NARCIS (Netherlands)

    Nazaretian, S.P.; Schenberg, M.E.; Simpson, I.; Slootweg, P.J.

    2007-01-01

    Ghost cell odontogenic carcinoma (GCOC) is the malignant counterpart of calcifying cystic odontogenic tumour and dentinogenic ghost cell tumour. This is the case of a middle-aged male who presented with a slow-growing maxillary tumour. He was asymptomatic until pain symptoms developed prior to initi

  5. Subungual squamous cell carcinoma*

    Science.gov (United States)

    Padilha, Carolina Barbosa de Sousa; Balassiano, Laila Klotz de Almeida; Pinto, Julyana Calegari; de Souza, Flávia Crespo Schueler; Kac, Bernard Kawa; Treu, Curt Mafra

    2016-01-01

    Although subungual squamous cell carcinoma is rare, it is the most common primary malignant neoplasms in this location. The higher incidence occurs in the fingernails, but involvement of the toenails is also possible. Subungual squamous cell carcinoma often looks like other more common benign lesions, such as fungal infection, onychomycosis, or viral wart. These factors, together with a general lack of awareness of this disease among physicians, often result in delayed diagnosis. Therefore, it is underdiagnosed, with few reports in the literature. The authors present a case of a man with a diagnosis of subungual squamous cell carcinoma in the hallux, without bone involvement, which was submitted to the appropriate surgical treatment. PMID:28099608

  6. Retraction: "Down-regulation of Notch-1 is associated with Akt and FoxM1 in inducing cell growth inhibition and apoptosis in prostate cancer cells" by Wang et al.

    Science.gov (United States)

    2016-08-01

    The above article, published online on July 23, 2010 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the journal Editor in Chief, Gary S. Stein, and Wiley Periodicals, Inc. The retraction has been agreed following an investigation from Wayne State University involving the first author and the corresponding author that found Figures 1B and 3A to be inappropriately manipulated. REFERENCE Wang Z, Li Y, Ahmad A, Banerjee S, Azmi AS, Kong D, Wojewoda C, Miele L, Sarkar FH. 2011. Down-regulation of Notch-1 is associated with Akt and FoxM1 in inducing cell growth inhibition and apoptosis in prostate cancer cells. J Cell Biochem 112:78-88; doi: 10.1002/jcb.22770.

  7. Retraction: "Down-regulation of uPA and uPAR by 3,3'-diindolylmethane contributes to the inhibition of cell growth and migration of breast cancer cells" by Ahmad et al.

    Science.gov (United States)

    2016-08-01

    The above article, published online on August 19, 2009 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the journal Editor in Chief, Gary S. Stein, and Wiley Periodicals, Inc. The retraction has been agreed following an investigation from Wayne State University involving the third author and the corresponding author that found Figure 5C to be inappropriately re-used and re-labeled. REFERENCE Ahmad A, Kong D, Wang Z, Sarkar SH, Banerjee S, Sarkar FH. 2009. Down-regulation of uPA and uPAR by 3,3'-diindolylmethane contributes to the inhibition of cell growth and migration of breast cancer cells. J Cell Biochem 108:916-925; doi: 10.1002/jcb.22323.

  8. Hibiscus sabdariffa Leaf Extract Inhibits Human Prostate Cancer Cell Invasion via Down-Regulation of Akt/NF-kB/MMP-9 Pathway

    Directory of Open Access Journals (Sweden)

    Chun-Tang Chiu

    2015-06-01

    Full Text Available Hibiscus sabdariffa leaf has been previously shown to possess hypoglycemic, hypolipidemic, and antioxidant effects, and induce tumor cell apoptosis. However, the molecular mechanisms involved in the anticancer activity of H. sabdariffa leaf extract (HLE are poorly understood. The object of the study was to examine the anti-invasive potential of HLE. First, HLE was demonstrated to be rich in polyphenols. The results of wound-healing assay and in vitro transwell assay revealed that HLE dose-dependently inhibited the migration and invasion of human prostate cancer LNCaP (lymph node carcinoma of the prostate cells under non-cytotoxic concentrations. Our results further showed that HLE exerted an inhibitory effect on the activity and expressions of matrix metalloproteinase-9 (MMP-9. The HLE-inhibited MMP-9 expression appeared to be a consequence of nuclear factor-kappaB (NF-κB inactivation because its DNA-binding activity was suppressed by HLE. Molecular data showed all these influences of HLE might be mediated via inhibition of protein kinase B (PKB, also known as Akt/NF-kB/MMP-9 cascade pathway, as demonstrated by the transfection of Akt1 overexpression vector. Finally, the inhibitory effect of HLE was proven by its inhibition on the growth of LNCaP cells and the expressions of metastasis-related molecular proteins in vivo. These findings suggested that the inhibition of MMP-9 expression by HLE may act through the suppression of the Akt/NF-kB signaling pathway, which in turn led to the reduced invasiveness of the cancer cells.

  9. Down-regulation of TSC-22 (transforming growth factor beta-stimulated clone 22) markedly enhances the growth of a human salivary gland cancer cell line in vitro and in vivo.

    Science.gov (United States)

    Nakashiro, K; Kawamata, H; Hino, S; Uchida, D; Miwa, Y; Hamano, H; Omotehara, F; Yoshida, H; Sato, M

    1998-02-01

    We have recently isolated TSC-22 (transforming growth factor beta-stimulated clone 22) cDNA as a new anticancer drug (Vesnarinone)-inducible gene in a human salivary gland cancer cell line, TYS. We conducted the present study to examine whether up-regulation or down-regulation of TSC-22 can affect the growth of TYS cells in vitro and in vivo. We constructed an expression vector containing sense- or antisense-oriented human TSC-22 cDNA under the transcriptional control of the SR alpha promoter. We cotransfected TYS cells with the sense or antisense expression vector and pSV2neo and obtained more than 200 G418-resistant colonies in each sense or antisense transfectant. Approximately 80% of representative G418-resistant clones expressed the transcripts from transfected sense or antisense TSC-22 cDNA. To avoid the clonal heterogeneity of the cells, we mixed all of the G418-resistant colonies together in each sense or antisense transfectant and examined the expression of TSC-22 protein, in vitro growth, and the tumorigenicity in nude mice. The expression of TSC-22 protein was examined by solid-phase ELISA using a specific antibody against recombinant TSC-22 protein. The expression of TSC-22 protein was up-regulated in the sense transfectants and down-regulated in the antisense transfectants. Contrary to our expectation, up-regulation of TSC-22 protein did not affect both in vitro and in vivo growth of TYS cells. However, down-regulation of TSC-22 markedly enhanced the growth of TYS cells in vitro and in vivo. Furthermore, we examined the expression of TSC-22 mRNA in several human salivary gland tumors. The mRNA expression of TSC-22 in benign and malignant salivary gland tumors was significantly decreased when compared to that in tumor-free salivary glands (P way ANOVA), and in some salivary gland tumors, the expression of TSC-22 mRNA was not detectable by reverse transcription-PCR. These results suggest that down-regulation of TSC-22 may play a major role on salivary

  10. Down-Regulation of Neurocan Expression in Reactive Astrocytes Promotes Axonal Regeneration and Facilitates the Neurorestorative Effects of Bone Marrow Stromal Cells in the Ischemic Rat Brain

    Institute of Scientific and Technical Information of China (English)

    LI HONG SHEN; YI LI; QI GAO; SMITA SAVANT-BHONSALE; MICHAEL CHOPP

    2008-01-01

    cerebral artery occlusion followed by an injection of 3×106 rat BMSCs (n=16) or phosphate-buffered saline (n=15) into the tail vein 24 h later. Animals were sacrificed at 8 days after stroke. Immunostaining analysis showed that reactive astrocytes were the primary source of neurocan, and BMSC-treated animals had significantly lower neurocan and higher growth associated protein 43 expression in the penumbral region compared with control rats, which was confirmed by Western blot analysis of the brain tissue. To further investigate the effects of BMSCs on astrocyte neurocan expression, single reactive astrocytes were collected from the ischemic boundary zone using laser capture microdissection. Neurocan gene expression was significantly down-regulated in rats receiving BMSC transplantation (n=4/group). Primary cultured astrocytes showed similar alterations; BMSC coculture during reoxygenation abolished the up-regulation of neurocan gene in astrocytes undergoing oxygen-glucose deprivation (n=3/group). Our data suggest that BMSCs promote axonal regeneration by reducing neurocan expression in peri-infarct astrocytes.c 2008 Wiley-Liss,Inc.

  11. c-myc down-regulation induces apoptosis in human cancer cell lines exposed to RPR-115135 (C31H29NO4), a non-peptidomimetic farnesyltransferase inhibitor.

    Science.gov (United States)

    Russo, Patrizia; Arzani, Dario; Trombino, Sonya; Falugi, Carla

    2003-01-01

    A therapeutic strategy that relies on the use of c-myc antisense in combination with a farnesyltransferase inhibitor, RPR-115135 (C31H29NO4), was studied in human cancer cell lines carrying different mutations (Ras, p53, myc amplification). Cell proliferation was strongly inhibited by the combination and was observed when c-myc oligo (at a concentration that down-regulates c-myc expression) was followed by RPR-115135. Cell cycle analysis demonstrated an accumulation in G0-G1 phase and a tendency to apoptosis (not detectable in cells treated with a single agent). Morphological examination and DNA fragmentation assays (filter binding and enzyme-linked immunosorbent assay DNA fragmentation) confirmed the induction of apoptosis. Apoptosis was not p53- and/or p21(waf-1)-dependent, and the key effector was caspase activation. The combination induced Bax expression and Bcl-2 inhibition. Down-regulation of c-myc amplification carried out a specific role exclusively when Ras was mutated. Exposure of human proliferating lymphocytes to combination did not result in cytotoxicity, suggesting that mechanisms regulating c-myc gene expression during normal T cell proliferation might not be involved. Because of the high percentage of human tumors overexpressing c-myc mRNA and/or protein and, simultaneously, harboring oncogenic Ras mutants (i.e., colon cancers), interrupting the myc- and Ras-signaling pathway would be one of the major focuses on therapy of these types of tumors.

  12. Extrapulmonary small cell carcinoma.

    NARCIS (Netherlands)

    Heijden, E. van der; Heijdra, Y.F.

    2005-01-01

    This article reviews the recent literature on extrapulmonary small cell carcinomas. Until now, only four cases have been published in the English literature, two of those in the Southern Medical Journal. Sharing the information on diagnosis and treatment of these cases is important for better unders

  13. Suppression of c-Myc is involved in multi-walled carbon nanotubes' down-regulation of ATP-binding cassette transporters in human colon adenocarcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Zhaojing [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China); Xu, Yonghong [Institute of Ophthalmological Research, Department of Ophthalmology, Renmin Hospital of Wuhan University, 430060 Wuhan (China); Meng, Xiangning [School of Materials and Metallurgy, Northeastern University, Shenyang 110819 (China); Watari, Fumio [Department of Biomedical, Dental Materials and Engineering, Graduate School of Dental Medicine, Hokkaido University, Sapporo 060-8586 (Japan); Liu, Hudan, E-mail: hudanliu@hust.edu.cn [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China); Chen, Xiao, E-mail: mornsmile@yahoo.com [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China)

    2015-01-01

    Over-expression of ATP-binding cassette (ABC) transporters, a large family of integral membrane proteins that decrease cellular drug uptake and accumulation by active extrusion, is one of the major causes of cancer multi-drug resistance (MDR) that frequently leads to failure of chemotherapy. Carbon nanotubes (CNTs)-based drug delivery devices hold great promise in enhancing the efficacy of cancer chemotherapy. However, CNTs' effects on the ABC transporters remain under-investigated. In this study, we found that multiwalled carbon nanotubes (MWCNTs) reduced transport activity and expression of ABC transporters including ABCB1/Pgp and ABCC4/MRP4 in human colon adenocarcinoma Caco-2 cells. Proto-oncogene c-Myc, which directly regulates ABC gene expression, was concurrently decreased in MWCNT-treated cells and forced over-expression of c-Myc reversed MWCNTs' inhibitory effects on ABCB1 and ABCC4 expression. MWCNT-cell membrane interaction and cell membrane oxidative damage were observed. However, antioxidants such as vitamin C, β-mecaptoethanol and dimethylthiourea failed to antagonize MWCNTs' down-regulation of ABC transporters. These data suggest that MWCNTs may act on c-Myc, but not through oxidative stress, to down-regulate ABC transporter expression. Our findings thus shed light on CNTs' novel cellular effects that may be utilized to develop CNTs-based drug delivery devices to overcome ABC transporter-mediated cancer chemoresistance.

  14. CD4(+) T-cell activation is differentially modulated by bacteria-primed dendritic cells, but is generally down-regulated by n-3 polyunsaturated fatty acids.

    Science.gov (United States)

    Brix, Susanne; Lund, Pia; Kjaer, Tanja M R; Straarup, Ellen M; Hellgren, Lars I; Frøkiaer, Hanne

    2010-03-01

    Appropriate activation of CD4(+) T cells is fundamental for efficient initiation and progression of acquired immune responses. Here, we showed that CD4(+) T-cell activation is dependent on changes in membrane n-3 polyunsaturated fatty acids (PUFAs) and is dynamically regulated by the type of signals provided by dendritic cells (DCs). Upon interaction with DCs primed by different concentrations and species of gut bacteria, CD4(+) T cells were activated according to the type of DC stimulus. The levels of CD80 were found to correlate to the levels of expression of CD28 and to the proliferation of CD4(+) T cells, while the presence of CD40 and CD86 on DCs inversely affected inducible costimulator (ICOS) and cytotoxic T-lymphocyte antigen-4 (CTLA-4) levels in CD4(+) T cells. For all DC stimuli, cells high in n-3 PUFAs showed reduced ability to respond to CD28 stimulation, to proliferate, and to express ICOS and CTLA-4. Diminished T-cell receptor (TCR) and CD28 signalling was found to be responsible for n-3 PUFA effects. Thus, the dietary fatty acid composition influences the overall level of CD4(+) T-cell activation induced by DCs, while the priming effect of the DC stimuli modulates CD80, CD86 and CD40 levels, thereby affecting and shaping activation of acquired immunity by differential regulation of proliferation and costimulatory molecule expression in CD4(+) T cells.

  15. Lack of estrogen down-regulates CXCR4 expression on Treg cells and reduces Treg cell population in bone marrow in OVX mice.

    Science.gov (United States)

    Fan, X-L; Duan, X-B; Chen, Z-H; Li, M; Xu, J-S; Ding, G-M

    2015-05-08

    Postmenopausal osteoporosis (PMO) is the most common metabolic bone disease in women after menopausal. Recent works focused on cross—talk between immune regulation and bone metabolism pathways and suggested Treg cells suppressed bone resorption and osteoclasts (OC) differentiation in bone marrow via cell—cell contact interaction and/or secreting of IL—10 and TGF—beta. In this study, we investigated the impact of estrogen on regulatory T cells (Treg cells) trafficking and staying in bone marrow and we found that a significant reduction of Treg cell population in bone marrow in estrogen deficiency ovariectomied (OVX) mice. We then studied the expressions of chemokines CXCL12/CXCR4 axes, which were critical to Treg cells migration and our data show the expression of CXCR4 on Treg cells was relative with oestrogen in vivo, however, the expression of CXCL12 was not. Furthermore, the loss of trafficking ability of Treg cells in OVX mice was recoverable in our system. These findings may mechanistically explain why Treg cells lose their suppressive functions on the regulation of OC cells and demonstrate a previously unappreciated role for estrogen, which may be critical to the novel therapy in clinical practice of PMO patients.

  16. Enriched environment inhibits mouse pancreatic cancer growth and down-regulates the expression of mitochondria-related genes in cancer cells.

    Science.gov (United States)

    Li, Guohua; Gan, Yu; Fan, Yingchao; Wu, Yufeng; Lin, Hechun; Song, Yanfang; Cai, Xiaojin; Yu, Xiang; Pan, Weihong; Yao, Ming; Gu, Jianren; Tu, Hong

    2015-01-19

    Psycho-social stress has been suggested to influence the development of cancer, but it remains poorly defined with regard to pancreatic cancer, a lethal malignancy with few effective treatment modalities. In this study, we sought to investigate the impacts of enriched environment (EE) housing, a rodent model of "eustress", on the growth of mouse pancreatic cancer, and to explore the potential underlying mechanisms through gene expression profiling. The EE mice showed significantly reduced tumor weights in both subcutaneous (53%) and orthotopic (41%) models, while each single component of EE (inanimate stimulation, social stimulation or physical exercise) was not profound enough to achieve comparative anti-tumor effects as EE. The integrative transcriptomic and proteomic analysis revealed that in response to EE, a total of 129 genes in the tumors showed differential expression at both the mRNA and protein levels. The differentially expressed genes were mostly localized to the mitochondria and enriched in the citrate cycle and oxidative phosphorylation pathways. Interestingly, nearly all of the mitochondria-related genes were down-regulated by EE. Our data have provided experimental evidence in favor of the application of positive stress or of benign environmental stimulation in pancreatic cancer therapy.

  17. Maternal immune stimulation reduces both placental morphologic damage and down-regulated placental growth-factor and cell cycle gene expression caused by urethane: are these events related to reduced teratogenesis?

    Science.gov (United States)

    Sharova, L V; Sharov, A A; Sura, P; Gogal, R M; Smith, B J; Holladay, S D

    2003-07-01

    Activation of the maternal immune system in mice decreased cleft palate caused by the chemical teratogen, urethane. Direct and indirect mechanisms for this phenomenon have been suggested, including maternal macrophages that cross the placenta to find and eliminate pre-teratogenic cells, or maternal immune proteins (cytokines) that cross placenta to alleviate or partially alleviate toxicant-mediated effects in the developing fetus. A third mechanism to explain improved fetal developmental outcome in teratogen-challenged pregnant mice might involve beneficial effects of immune stimulation on the placenta. In the present experiments, urethane treatment altered placental morphology and impaired placental function, the latter indicated by down-regulated activity of cell cycle genes and of genes encoding cytokines and growth factors. Maternal immune stimulation with either Freund's complete adjuvant (FCA) or interferon-gamma (IFNgamma) reduced morphologic damage to the placenta caused by urethane and normalized expression of several genes that were down-regulated by urethane. Urethane treatment also shifted placental cytokine gene expression toward a T cell helper 1 (Th1) profile, while immunostimulation tended to restore a Th2 profile that may be more beneficial to pregnancy and fetal development. These data suggest that the beneficial effects of maternal immune stimulation on fetal development in teratogen-exposed mice may, in part, result from improved placental structure and function.

  18. Human TM9SF4 Is a New Gene Down-Regulated by Hypoxia and Involved in Cell Adhesion of Leukemic Cells.

    Directory of Open Access Journals (Sweden)

    Rosa Paolillo

    Full Text Available The transmembrane 9 superfamily protein member 4, TM9SF4, belongs to the TM9SF family of proteins highly conserved through evolution. TM9SF4 homologs, previously identified in many different species, were mainly involved in cellular adhesion, innate immunity and phagocytosis. In human, the function and biological significance of TM9SF4 are currently under investigation. However, TM9SF4 was found overexpressed in human metastatic melanoma and in a small subset of acute myeloid leukemia (AMLs and myelodysplastic syndromes, consistent with an oncogenic function of this gene.In this study, we first analyzed the expression and regulation of TM9SF4 in normal and leukemic cells and identified TM9SF4 as a gene highly expressed in human quiescent CD34+ hematopoietic progenitor cells (HPCs, regulated during monocytic and granulocytic differentiation of HPCs, both lineages giving rise to mature myeloid cells involved in adhesion, phagocytosis and immunity. Then, we found that TM9SF4 is markedly overexpressed in leukemic cells and in AMLs, particularly in M2, M3 and M4 AMLs (i.e., in AMLs characterized by the presence of a more or less differentiated granulocytic progeny, as compared to normal CD34+ HPCs. Proliferation and differentiation of HPCs occurs in hypoxia, a physiological condition in bone marrow, but also a crucial component of cancer microenvironment. Here, we investigated the impact of hypoxia on TM9SF4 expression in leukemic cells and identified TM9SF4 as a direct target of HIF-1α, downregulated in these cells by hypoxia. Then, we found that the hypoxia-mediated downregulation of TM9SF4 expression is associated with a decrease of cell adhesion of leukemic cells to fibronectin, thus demonstrating that human TM9SF4 is a new molecule involved in leukemic cell adhesion.Altogether, our study reports for the first time the expression of TM9SF4 at the level of normal and leukemic hematopoietic cells and its marked expression at the level of AMLs

  19. Bortezomib and Fenretinide Induce Synergistic Cytotoxicity in Mantle Cell Lymphoma Via Apoptosis, Cell Cycle Dysregulation, and IκBα Kinase Down-regulation

    Science.gov (United States)

    Cowan, Andrew J.; Frayo, Shani L.; Press, Oliver W.; Palanca-Wessels, Maria C.; Pagel, John M.; Green, Damian J.; Gopal, Ajay K.

    2015-01-01

    Background Mantle cell lymphoma (MCL) remains incurable for most patients and proteasome inhibitors like bortezomib induce responses in a minority of patients with relapsed disease. Fenretinide is a retinoid that has shown preclinical activity in B-cell lymphomas. We hypothesized that these agents could yield augmented anti-tumor activity. Methods Mantle cell lymphoma lines (Granta-519, Jeko-1, Rec-1) were treated with escalating concentrations of bortezomib and fenretinide singly and in combination. Cytotoxicity was assessed using the MTT assay. Flow cytometric methods assessed apoptosis and necrosis with annexin V-FITC/propidium iodide and G1 and G2 cell cycle changes with DAPI staining. Changes in Cyclin D1, Cyclin B, IκBα, and IKKα expression were quantified by Western blot. Results Cytotoxicity was mediated via apoptosis; both agents showed observed vs expected cytotoxicity in Granta-519 of 92.2% vs 55.1%, in Jeko-1 of 87.6% vs 36.3%, and in Rec-1 of 63.2% vs 29.8%. Isobolographic analysis confirmed synergy in Jeko-1 and Rec-1. Bortezomib induced G2 phase arrest with a 1.7 fold-increase over control, and fenretinide resulted in G1 phase arrest, with an increase of 1.3 fold over control. In combination G2 phase arrest predominated, with a 1.4 fold-increase compared to control, and reduced expression of Cyclin D1 to 24%, Cyclin B to 52% and 64%, Cyclin D3 to 25% and 43%, IκBα to 23% and 46%, and IκBα kinase to 34% and 44%. Conclusions Bortezomib and fenretinide exhibit synergistic cytotoxicity against MCL cell lines. This activity is mediated by IκBα kinase modulation, decreased cyclin expression, cell cycle dysregulation, and apoptotic cell death. PMID:26237500

  20. Down-regulation of c9orf86 in human breast cancer cells inhibits cell proliferation, invasion and tumor growth and correlates with survival of breast cancer patients.

    Directory of Open Access Journals (Sweden)

    Yang-Yang Li

    Full Text Available C9orf86 which is a novel subfamily within the Ras superfamily of GTPases, is overexpressed in the majority of primary breast tumors. Few functional studies have focused on the C9orf86 protein; therefore, in this study, we explored the role of C9orf86 in breast carcinogenesis. In our study, we found that silencing of C9orf86 by siRNA in MCF-7 and SK-BR-3 cells resulted in suppressed cell proliferation as well as in vitro cell invasion capabilities. Moreover, knockdown of C9orf86 inhibited tumor growth in nude mice. Cell cycle and apoptotic assays showed that the anti-proliferative effect of C9orf86-siRNA was mediated by arresting cells in the G1 phase and promoting apoptosis. In addition, we found that patients with high levels of C9orf86 expression showed a significant trend towards worse survival compared to patients with low C9orf86 expression (P = 0.002. These results provide evidence that C9orf86 represents a novel and clinically useful biomarker for BC patients and plays an important role during the progression of BC.

  1. Human cytomegalovirus-encoded miR-US4-1 promotes cell apoptosis and benefits discharge of infectious virus particles via down-regulation of glutaminyl-tRNA synthetase, QARS in HCMV-infected HELF cells

    Indian Academy of Sciences (India)

    Yaozhong Shao; Ying Qi; Yujing Huang; Zhongyang Liu; Yanping Ma; Xin Guo; Shujuan Jiang; Zhengrong Sun; Qiang Ruan

    2016-06-01

    Human cytomegalovirus (HCMV) can cause congenital diseases and opportunistic infections in immunocompromised individuals. Its functional proteins and microRNAs (miRNAs) facilitate efficient viral propagation by altering host cell behaviour. Identification of functional target genes of miRNAs is an important step in studies on HCMV pathogenesis. In this study, Glutaminyl-tRNA Synthetase (QARS), which could regulate signal transduction pathways for cellular apoptosis, was identified as a direct target of hcmv-miR-US4-1. Apoptosis assay revealed that as silence of QARS by ectopic expression of hcmv-miR-US4-1 and specific small interference RNA of QARS can promote cell apoptosis in HCMV-infected HELF cells. Moreover, viral growth curve assays showed that hcmv-miR-US4-1 benefits the discharge of infectious virus particles. However, silence of hcmv-miR-US4-1 by its specific inhibitor overturned these effects. These results imply that hcmv-miR-US4-1 might have the same effects during HCMV nature infection. In general, hcmv-miR-US4-1 may involve in promoting cell apoptosis and benefiting discharge of infectious virus particles via down-regulation of QARS in HCMV-infected HELF cells.

  2. HHV-8 reduces dendritic cell migration through down-regulation of cell-surface CCR6 and CCR7 and cytoskeleton reorganization

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    Cirone Mara

    2012-05-01

    Full Text Available Abstract Background For an efficient immune response against viral infection, dendritic cells (DCs must express a coordinate repertoire of receptors that allow their recruitment to the sites of inflammation and subsequently to the secondary lymphoid organs in response to chemokine gradients. Several pathogens are able to subvert the chemokine receptor expression and alter the migration properties of DCs as strategy to escape from the immune control. Findings Here we report the inhibitory effect of Human Herpesvirus 8 (HHV-8 on the migratory behavior of immature and mature DCs. We found that the virus altered the DC chemokine receptor expression and chemokine induced migration. Moreover HHV-8 was also able to interfere with basal motility of DCs by inducing cytoskeleton modifications. Conclusion Based on our findings, we suggest that HHV-8 is able to subvert the DC migration capacity and this represents an additional mechanism which interferes with their immune-functions.

  3. Acute cold exposure-induced down-regulation of CIDEA, cell death-inducing DNA fragmentation factor-alpha-like effector A, in rat interscapular brown adipose tissue by sympathetically activated beta3-adrenoreceptors.

    Science.gov (United States)

    Shimizu, Takahiro; Yokotani, Kunihiko

    2009-09-18

    The thermogenic activity of brown adipose tissue (BAT) largely depends on the mitochondrial uncoupling protein 1 (UCP1), which is up-regulated by environmental alterations such as cold. Recently, CIDEA (cell death-inducing DNA fragmentation factor-alpha-like effector A) has also been shown to be expressed at high levels in the mitochondria of BAT. Here we examined the effect of cold on the mRNA and protein levels of CIDEA in interscapular BAT of conscious rats with regard to the sympathetic nervous system. Cold exposure (4 degrees C for 3h) elevated the plasma norepinephrine level and increased norepinephrine turnover in BAT. Cold exposure resulted in down-regulation of the mRNA and protein levels of CIDEA in BAT, accompanied by up-regulation of mRNA and protein levels of UCP1. The cold exposure-induced changes of CIDEA and UCP1 were attenuated by intraperitoneal pretreatment with propranolol (a non-selective beta-adrenoreceptor antagonist) (2mg/animal) or SR59230A (a selective beta(3)-adrenoreceptor antagonist) (2mg/animal), respectively. These results suggest that acute cold exposure resulted in down-regulation of CIDEA in interscapular BAT by sympathetically activated beta(3)-adrenoreceptor-mediated mechanisms in rats.

  4. The GATA1s isoform is normally down-regulated during terminal haematopoietic differentiation and over-expression leads to failure to repress MYB, CCND2 and SKI during erythroid differentiation of K562 cells

    Directory of Open Access Journals (Sweden)

    Halsey Christina

    2012-08-01

    Full Text Available Abstract Background Although GATA1 is one of the most extensively studied haematopoietic transcription factors little is currently known about the physiological functions of its naturally occurring isoforms GATA1s and GATA1FL in humans—particularly whether the isoforms have distinct roles in different lineages and whether they have non-redundant roles in haematopoietic differentiation. As well as being of general interest to understanding of haematopoiesis, GATA1 isoform biology is important for children with Down syndrome associated acute megakaryoblastic leukaemia (DS-AMKL where GATA1FL mutations are an essential driver for disease pathogenesis. Methods Human primary cells and cell lines were analyzed using GATA1 isoform specific PCR. K562 cells expressing GATA1s or GATA1FL transgenes were used to model the effects of the two isoforms on in vitro haematopoietic differentiation. Results We found no evidence for lineage specific use of GATA1 isoforms; however GATA1s transcripts, but not GATA1FL transcripts, are down-regulated during in vitro induction of terminal megakaryocytic and erythroid differentiation in the cell line K562. In addition, transgenic K562-GATA1s and K562-GATA1FL cells have distinct gene expression profiles both in steady state and during terminal erythroid differentiation, with GATA1s expression characterised by lack of repression of MYB, CCND2 and SKI. Conclusions These findings support the theory that the GATA1s isoform plays a role in the maintenance of proliferative multipotent megakaryocyte-erythroid precursor cells and must be down-regulated prior to terminal differentiation. In addition our data suggest that SKI may be a potential therapeutic target for the treatment of children with DS-AMKL.

  5. 下调βIII-tubulin逆转肺腺癌A549/Taxol细胞株紫杉醇耐药%Down-regulatedβIII-tubulin Expression Can Reverse Paclitaxel Resistance in A549/Taxol Cells Lines

    Institute of Scientific and Technical Information of China (English)

    禚银玲; 郭其森

    2014-01-01

    Background and objective Chemotherapy drug resistance is the primary causes of death in patients with pulmonary carcinoma which make tumor recurrence or metastasis.β-tubulin is the main cell targets of anti-microtubule drug. Increased expression ofβIII-tubulin has been implicated in non-small cell lung cancer (NSCLC) cell lines. To explore the relationship among the expression level ofβIII-tubulin and the sensitivity of A549/Taxolcell lines to Taxol and cell cycles and cell apoptosis by RNA interference-mediated inhibition ofβIII-tubulin in A549/Taxol cells. Methods hTree pairs of siRNA targetdβIII-tubulin were designed and prepared, which were transfected into A549/Taxol cells using LipofectamineTM 2000. We detected the expression ofβIII-tubulin mRNA using Real-time lfuorescence qRT-PCR. Tedhen we selected the most eff-cient siRNA by the expression ofβIII-tubulin mRNA in transfected group.βIII-tubulin protein level were mesured by Western blot. hTe taxol sensitivity in transfected group were evaluated by MTT assay. And the cell apoptosis and cell cycles were deter-mined by lfow cytometry. Results βIII-tubulin mRNA levels in A549/Taxol cells were signiifcantly decreased in transfected grop by Real-time qRT-PCR than control groups. AndβIII-tubulin siRNA-1 sequence showed the highest transfection eff-ciency, which was (87.73±4.87)%(P<0.01);Western blot results showed that the expressional level of BIII tublin protein was signiifcantly down-reulated in the transfectant cells than thant in the control cells. By MTT assay, we showed that the inhibition ratio of Taxol to A549/Taxol cells transfeced was higher than that of control group (51.77±4.60)%(P<0.01). hTe early apopto-sis rate of A549/Taxol cells in transfected group were signiifcantly higher than that of control group (P<0.01);G2-M content in taxol group obviously increased than untreated samples by the cell cycle (P<0.05). Conclusion βIII-tubulin down-regulated signiifcantly sensitized NSCLC A549

  6. Is telomerase reactivation associated with the down-regulation of TGF β receptor-II expression in human breast cancer?

    Directory of Open Access Journals (Sweden)

    Thomas Valene

    2003-07-01

    Full Text Available Abstract Background Telomerase is a ribonucleoprotein that synthesizes telomeres and plays an important role in chromosomal stability and cellular immortalisation. Telomerase activity is detectable in most human cancers but not in normal somatic cells. TGF beta (transforming growth factor beta is a member of a family of cytokines that are essential for cell survival and seems to be down-regulated in human cancer. Recent in vitro work using human breast cancer cell lines has suggested that TGF beta down-regulates the expression of hTERT (human telomerase reverse transcriptase : the catalytic subunit of telomerase. We have therefore hypothesised that telomerase reactivation is associated with reduced immunohisto-chemical expression of TGF beta type II receptor (RII in human breast cancer. Methods TGF beta RII immunohistochemical expression was determined in 24 infiltrating breast carcinomas with known telomerase activity (17 telomerase-positive and 7 telomerase-negative. Immunohistochemical expression of TGF beta RII was determined by a breast pathologist who was blinded to telomerase data. Results TGF beta RII was detected in all lesions. The percentage of stained cells ranged from 1–100%. The difference in TGF beta RII expression between telomerase positive and negative tumours was not statistically significant (p = 1.0. Conclusion The results of this pilot study suggest that there is no significant association between telomerase reactivation and TGF-beta RII down-regulation in human breast cancer.

  7. Retraction: "Down-regulation of Notch-1 and Jagged-1 inhibits prostate cancer cell growth, migration and invasion, and induces apoptosis via inactivation of Akt, mTOR, and NF-κB signaling pathways" by Wang et al.

    Science.gov (United States)

    2016-08-01

    The above article, published online on January 5, 2010 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the journal Editor in Chief, Gary S. Stein, and Wiley Periodicals, Inc. The retraction has been agreed following an investigation from Wayne State University involving the first author and the corresponding author that found Figure 5A to be inappropriately manipulated. REFERENCE Wang Z, Li Y, Banerjee S, Kong D, Ahmad A, Nogueira V, Hay N, Sarkar FH. 2010. Down-regulation of Notch-1 and Jagged-1 inhibits prostate cancer cell growth, migration and invasion, and induces apoptosis via inactivation of Akt, mTOR, and NF-κB signaling pathways. J Cell Biochem 109:726-736; doi: 10.1002/jcb.22451.

  8. Vismodegib in basal cell carcinoma.

    Science.gov (United States)

    Amaria, R N; Bowles, D W; Lewis, K D; Jimeno, A

    2012-07-01

    Vismodegib is a novel, small-molecule inhibitor of smoothened, a key component of the hedgehog signaling pathway. Increased hedgehog pathway signaling is critical in the development of hereditary and spontaneous basal cell carcinomas of the skin, and has been implicated in the development of a number of other tumors. In preclinical models, vismodegib demonstrated potent antitumor activity in hedgehog-dependent tumors, particularly basal cell carcinomas. Clinically, phase I and II studies showed dramatic anticancer activity in patients with advanced basal cell carcinomas. In January 2012, vismodegib was approved by the FDA for the treatment of unresectable or metastatic basal cell carcinomas of the skin.

  9. Down-regulation of triose phosphate isomerase in Vineristine-resistant gastric cancer SGC7901 cell line identified by immobilized pH gradient two-dimensional gel electrophoresis and mierosequencing

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Objective:To exkplore new multidrug-resistance-related proteins in gastric SC7901 cells and clarify their mechanisms.Methods:Two-dimensional(2-D) polyacrylamide gel electrophoresis with immobilized pH gradients(IPG) was applied to compare the differential expression of multidrug-resistance-related proteins in gastric cancer SGC7901 cells and Vineristine-resistant SGC7901 cells (SGC7901/VCR) induced by vincristine sulfate.The 2-D gels were silver-stained.Then,preparative 2-D PAGE was performed.The differential proteins of PVDF membranes were cxcised and identified by N-terminal microsequencing.The mRNA expressions of differential proteins were detected in SGC 7901 cells and SGC7901/VCR cells by RT-PCR.Results:Approximatedly 680 protein sports were resolved on each 2-D gel by silver staining.Most protein spots showed no difference in composition,shape or density.25 proteins differed in abundance (6 higher in SGC7901/VCR cells;19 higher in 7901 cells);5 proteins were unique to one kind of cell or the othe(3 in SGC7901/VRC cells,2 in 7901 cells).One drug-resistance-related protein,which was down-regulated in SGC7901/VCR cells,was identified as trisephosphate isomerase(TPI),a glycolytic pathway enzyme.Conclusions:the results suggest that these differential proteins including TPI may be related to the Vincristine-resistant mechanism in human gastric cancer SGC7901/VCR cell line.

  10. Penis squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Leonor Hernández Piñero

    2015-09-01

    Full Text Available Cancer has become a first order health problem worldwide, despite the great diagnostic and therapeutic programs achieved during the last years. This is a clinical case of an 81- year-old patient with personal and social history of promiscuous and unprotected sexual behavior that shows a vegetative lesion in his gland and numerous inguinal adenopathies. Biopsy confirms the diagnosis of squamous cell carcinoma infiltrating the penis, which is a relatively rare pathology which is generally diagnosed belatedly. Partial amputation of the penis was considered to be performed, but there was no consent on behalf of his family. The patient’s general condition was getting worse until he died.

  11. Squamous cell carcinoma.

    Science.gov (United States)

    Webb, Julie L; Burns, Rachel E; Brown, Holly M; LeRoy, Bruce E; Kosarek, Carrie E

    2009-03-01

    Squamous cell carcinoma (SCC) is a relatively common, malignant neoplasm of dogs and cats that can arise in a variety of locations. The gross appearance of SCC can be variable and nonspecific, so definitive diagnosis requires microscopic examination of the tissue (cytology or histology). Several treatment modalities exist, but surgical excision, if possible, is regarded as the best treatment option. Early diagnosis and treatment of SCC are key because small, early-stage tumors are the most amenable to treatment and carry the best prognosis.

  12. Synergistic Effect of Subtoxic-dose Cisplatin and TRAIL to Mediate Apoptosis by Down-regulating Decoy Receptor 2 and Up-regulating Caspase-8, Caspase-9 and Bax Expression on NCI-H460 and A549 Cells

    Directory of Open Access Journals (Sweden)

    Xiaoyan Zhang

    2013-05-01

    Full Text Available Objective(s: Although tumor necrosis factor-related apoptosis-inducing ligand (TRAIL can selectively induce apoptosis in tumor cells, more than half of tumors including non-small cell lung cancer (NSCLC exhibit TRAIL-resistance. The purpose of this study was to determine whether subtoxic-dose cisplatin and TRAIL could synergistically enhance apoptosis on NSCLC cells and investigate its underlying mechanisms. Materials and Methods:NCI-H460 and A549 cells were treated with TRAIL alone, cisplatin alone or combination treatment in this study. The cytotoxicity was evaluated according to Sulforhodamine B assay, and apoptosis was examined using Hoechst 33342 staining and flow cytometry. The mRNA and protein levels of TRAIL receptors and apoptotic proteins including caspase-8, caspase-9, Bcl-2 and Bax were determined by RT-PCR and Western blotting, respectively. Results:Our results showed that NCI-H460 cells were sensitive to TRAIL, whereas A549 cells were resistant. However, subtoxic-dose cisplatin could enhance the both cells to TRAIL-mediated cell proliferation inhibition and apoptosis. The underlying mechanisms might be associated with the down-regulation of DcR2 and up-regulation of Caspase-8, Caspase-9 and Bax. Conclusion:Subtoxic-dose cisplatin could enhance both TRAIL- sensitive and TRAIL- resistant NSCLC cells to TRAIL-mediated apoptosis. These findings motivated further studies to evaluate such a combinatory therapeutic strategy against NSCLC in the animal models.

  13. Clear cell papillary renal cell carcinoma: micro-RNA expression profiling and comparison with clear cell renal cell carcinoma and papillary renal cell carcinoma.

    Science.gov (United States)

    Munari, Enrico; Marchionni, Luigi; Chitre, Apurva; Hayashi, Masamichi; Martignoni, Guido; Brunelli, Matteo; Gobbo, Stefano; Argani, Pedram; Allaf, Mohamad; Hoque, Mohammad O; Netto, George J

    2014-06-01

    Clear cell papillary renal cell carcinoma (CCPRCC) is a low-grade renal neoplasm with morphological characteristics mimicking both clear cell renal cell carcinoma (CCRCC) and papillary renal cell carcinoma (PRCC). However, despite some overlapping features, their morphological, immunohistochemical, and molecular profiles are distinct. Micro-RNAs (miRNAs) are small noncoding RNAs that play a crucial role in regulating gene expression and are involved in various biological processes, including cancer development. To better understand the biology of this tumor, we aimed to analyze the miRNA expression profile of a set of CCPRCC using microarray and quantitative reverse transcription-polymerase chain reaction. A total of 15 cases diagnosed as CCPRCC were used in this study. Among the most differentially expressed miRNA in CCPRCC, we found miR-210, miR-122, miR-34a, miR-21, miR-34b*, and miR-489 to be up-regulated, whereas miR-4284, miR-1202, miR-135a, miR-1973, and miR-204 were down-regulated compared with normal renal parenchyma. To identify consensus of differentially regulated miRNA between CCPRCC, CCRCC, and PRCC, we additionally determined differential miRNA expression using 2 publically available microarray data sets from the NCBI Gene Expression Omnibus database (GSE41282 and GSE3798). This comparison revealed that the miRNA expression profile of CCPRCC shows some overlapping characteristics between CCRCC and PRCC. Moreover, CCPRCC lacks dysregulation of important miRNAs typically associated with aggressive behavior. In summary, we describe the miRNA expression profile of a relatively infrequent type of renal cancer. Our results may help in understanding the molecular underpinning of this newly recognized entity.

  14. Pantethine Alters Lipid Composition and Cholesterol Content of Membrane Rafts, With Down-Regulation of CXCL12-Induced T Cell Migration.

    Science.gov (United States)

    van Gijsel-Bonnello, Manuel; Acar, Niyazi; Molino, Yves; Bretillon, Lionel; Khrestchatisky, Michel; de Reggi, Max; Gharib, Bouchra

    2015-10-01

    Pantethine, a natural low-molecular-weight thiol, shows a broad activity in a large range of essential cellular pathways. It has been long known as a hypolipidemic and hypocholesterolemic agent. We have recently shown that it exerts a neuroprotective action in mouse models of cerebral malaria and Parkinson's disease through multiple mechanisms. In the present study, we looked at its effects on membrane lipid rafts that serve as platforms for molecules engaged in cell activity, therefore providing a target against inappropriate cell response leading to a chronic inflammation. We found that pantethine-treated cells showed a significant change in raft fatty acid composition and cholesterol content, with ultimate downregulation of cell adhesion, CXCL12-driven chemotaxis, and transendothelial migration of various T cell types, including human Jurkat cell line and circulating effector T cells. The mechanisms involved include the alteration of the following: (i) CXCL12 binding to its target cells; (ii) membrane dynamics of CXCR4 and CXCR7, the two CXCL12 receptors; and (iii) cell redox status, a crucial determinant in the regulation of the chemokine system. In addition, we considered the linker for activation of T cells molecule to show that pantethine effects were associated with the displacement from the rafts of the acylated signaling molecules which had their palmitoylation level reduced.. In conclusion, the results presented here, together with previously published findings, indicate that due to its pleiotropic action, pantethine can downregulate the multifaceted process leading to pathogenic T cell activation and migration.

  15. Lycium chinensis Mill attenuates glutamate induced oxidative toxicity in PC12 cells by increasing antioxidant defense enzymes and down regulating ROS and Ca(2+) generation.

    Science.gov (United States)

    Olatunji, Opeyemi J; Chen, Hongxia; Zhou, Yifeng

    2016-03-11

    Lycium chinensis Mill is a famous traditional Chinese medicine which displays several medicinal activities including antioxidant and neuroprotective activities. However, the mechanism of action towards the neuroprotective action has not been fully elucidated. This work was aimed at investigating the neuroprotective effects of L. chinensis Mill against glutamate-induced oxidative neurotoxicity in PC12 cells. Oxidative cell death was induced with 5mM glutamate in PC12 cells. Cell viability, LDH release, intracellular Ca(2+) concentration, reactive oxygen species (ROS) accumulation, GSH-Px, CAT and SOD antioxidant enzyme levels were measured. Our results indicated that pretreatment of PC12 cells with L. chinensis Mill extracts markedly attenuated the loss of cell viability, the release of lactate dehydrogenase (LDH), Ca(2+) overload, ROS generation, and cell apoptosis induced by glutamate toxicity. Furthermore, L. chinensis Mill extracts also significantly increased the levels of innate antioxidant enzymes GSH-Px, SOD and CAT in glutamate-induced PC12 cells. Conclusively, our results provided substantial evidence that L. chinensis Mill protected PC12 cells against glutamate-induced cell death by attenuating ROS generation, Ca(2+) influx, and increased the antioxidant defense capacity of PC12 cells against oxidative stress damages, suggesting the possible potential of extracts from the plant as sources of bioactive molecules in the treatment of neurodegenerative disorders.

  16. Ginsenoside PPD’s Antitumor Effect via Down-Regulation of mTOR Revealed by Super-Resolution Imaging

    Directory of Open Access Journals (Sweden)

    Bo Teng

    2017-03-01

    Full Text Available Derived from Panax ginseng, the natural product 20(S-Protopanaxadiol (PPD has been reported for its cytotoxicity against several cancer cell lines. The molecular mechanism is, however, not well understood. Here we show that PPD significantly inhibits proliferation, induces apoptosis and causes G2/M cell cycle arrest in human laryngeal carcinoma cells (Hep-2 cells. PPD also decreases the levels of proteins related to cell proliferation. Moreover, PPD-induced apoptosis is characterized by a dose-dependent down-regulation of Bcl-2 expression and up-regulation of Bax, and is accompanied by the activation of Caspase-3 as well. Further molecular mechanism is revealed by direct stochastic optical reconstruction microscopy (dSTORM—a novel high-precision localization microscopy which enables effective resolution down to the order of 10 nm. It shows the expression and spatial arrangement of mTOR and its downstream effectors, demonstrating that this ginsenoside exerts its excellent anticancer effects via down-regulation of mTOR signaling pathway in Hep-2 cells. Taken together, our findings elucidate that the antitumor effect of PPD is associated with its regulation of mTOR expression and distribution, which encourages further studies of PPD as a promising therapeutic agent against laryngeal carcinoma.

  17. Establishment of an ovarian metastasis model and possible involvement of E-cadherin down-regulation in the metastasis.

    Science.gov (United States)

    Kuwabara, Yoshiko; Yamada, Taketo; Yamazaki, Ken; Du, Wen-Lin; Banno, Kouji; Aoki, Daisuke; Sakamoto, Michiie

    2008-10-01

    Clinical observations of cases of ovarian metastasis suggest that there may be a unique mechanism underlying ovarian-specific metastasis. This study was undertaken to establish an in vivo model of metastasis to the ovary, and to investigate the mechanism of ovarian-specific metastasis. We examined the capacity for ovarian metastasis in eight different human carcinoma cell lines by implantation in female NOD/SCID mice transvenously and intraperitoneally. By transvenous inoculation, only RERF-LC-AI, a poorly differentiated carcinoma cell line, frequently demonstrated ovarian metastasis. By intraperitoneal inoculation, four of the eight cell lines (HGC27, MKN-45, KATO-III, and RERF-LC-AI) metastasized to the ovary. We compared E-cadherin expression among ovarian metastatic cell lines and others. All of these four ovarian metastatic cell lines and HSKTC, a Krukenberg tumor cell line, showed E-cadherin down-regulation and others did not. E-cadherin was then forcibly expressed in RERF-LC-AI, and inhibited ovarian metastasis completely. The capacity for metastasizing to the other organs was not affected by E-cadherin expression. We also performed histological investigation of clinical ovarian-metastatic tumor cases. About half of all ovarian-metastatic tumor cases showed loss or reduction of E-cadherin expression. These data suggest that E-cadherin down-regulation may be involved in ovarian-specific metastasis.

  18. Interferon-β induced microRNA-129-5p down-regulates HPV-18 E6 and E7 viral gene expression by targeting SP1 in cervical cancer cells.

    Directory of Open Access Journals (Sweden)

    Jiarong Zhang

    Full Text Available Infection by human papillomavirus (HPV can cause cervical intraepithelial neoplasia (CIN and cancer. Down-regulation of E6 and E7 expression may be responsible for the positive clinical outcomes observed with IFN treatment, but the molecular basis has not been well determined. As miRNAs play an important role in HPV induced cervical carcinogenesis, we hypothesize that IFN-β can regulate the expressions of specific miRNAs in cervical cancer cells, and that these miRNAs can mediate E6 and E7 expression, thus modulate their oncogenic potential. In this study, we found that miR-129-5p to be a candidate IFN-β inducible miRNA. MiR-129-5p levels gradually decrease with the development of cervical intraepithelial lesions. Manipulation of miR-129-5p expression in Hela cells modulates HPV-18 E6 and E7 viral gene expression. Exogenous miR-129-5p inhibits cell proliferation in Hela cells, promotes apoptosis and blocks cell cycle progression in Hela cells. SP1 is a direct target of miR-129-5p in Hela cells. This study is the first report of a cellular miRNA with anti-HPV activity and provides new insights into regulatory mechanisms between the HPV and the IFN system in host cells at the miRNA level.

  19. PI3K/Akt inhibition and down-regulation of BCRP re-sensitize MCF7 breast cancer cell line to mitoxantrone chemotherapy

    Directory of Open Access Journals (Sweden)

    Tahereh Komeili-Movahhed

    2015-05-01

    Full Text Available Objective(s:Multidrug resistance (MDR of cancer cells is a major obstacle to successful chemotherapy. Overexpression of breast cancer resistance protein (BCRP is one of the major causes of MDR. In addition, it has been shown that PI3K/Akt signaling pathway involves in drug resistance. Therefore, we evaluated the effects of novel approaches including siRNA directed against BCRP and targeted therapy against PI3K/Akt signaling pathway using LY294002 (LY to re-sensitize breast cancer MCF7 cell line to mitoxantrone (MTX chemotherapy. Materials and Methods: Anticancer effects of MTX, siRNA, and LY alone and in combination were evaluated in MCF7 cells using MTT cytotoxicity assay and flow cytometry analysis of cell cycle distribution and apoptosis induction. Results: MTT and apoptosis assays showed that both MTX and LY inhibited cell proliferation and induced apoptosis in MCF7 cells. Results indicated that inhibition of BCRP by siRNA or PI3K/Akt signaling pathway by LY significantly increased sensitivity of MCF7 cells to antiproliferation and apoptosis induction of MTX. Furthermore, MTX showed G2/M arrest, whereas LY induced G0/G1 arrest in cell cycle distribution of MCF7 cells. Combination of siRNA or LY with MTX chemotherapy significantly increased accumulation of MCF7 cells in the G2/M phase of cell cycle. Conclusion: Combination of MTX chemotherapy with BCRP siRNA and PI3K/Akt inhibition can overcome MDR in breast cancer cells. This study furthermore suggests that novel therapeutic approaches are needed to enhance anticancer effects of available drugs in breast cancer

  20. PI3K/Akt inhibition and down-regulation of BCRP re-sensitize MCF7 breast cancer cell line to mitoxantrone chemotherapy

    Science.gov (United States)

    Komeili-Movahhed, Tahereh; Fouladdel, Shamileh; Barzegar, Elmira; Atashpour, Shekoufeh; Hossein Ghahremani, Mohammad; Nasser Ostad, Seyed; Madjd, Zahra; Azizi, Ebrahim

    2015-01-01

    Objective(s): Multidrug resistance (MDR) of cancer cells is a major obstacle to successful chemotherapy. Overexpression of breast cancer resistance protein (BCRP) is one of the major causes of MDR. In addition, it has been shown that PI3K/Akt signaling pathway involves in drug resistance. Therefore, we evaluated the effects of novel approaches including siRNA directed against BCRP and targeted therapy against PI3K/Akt signaling pathway using LY294002 (LY) to re-sensitize breast cancer MCF7 cell line to mitoxantrone (MTX) chemotherapy. Materials and Methods: Anticancer effects of MTX, siRNA, and LY alone and in combination were evaluated in MCF7 cells using MTT cytotoxicity assay and flow cytometry analysis of cell cycle distribution and apoptosis induction. Results: MTT and apoptosis assays showed that both MTX and LY inhibited cell proliferation and induced apoptosis in MCF7 cells. Results indicated that inhibition of BCRP by siRNA or PI3K/Akt signaling pathway by LY significantly increased sensitivity of MCF7 cells to antiproliferation and apoptosis induction of MTX. Furthermore, MTX showed G2/M arrest, whereas LY induced G0/G1 arrest in cell cycle distribution of MCF7 cells. Combination of siRNA or LY with MTX chemotherapy significantly increased accumulation of MCF7 cells in the G2/M phase of cell cycle. Conclusion: Combination of MTX chemotherapy with BCRP siRNA and PI3K/Akt inhibition can overcome MDR in breast cancer cells. This study furthermore suggests that novel therapeutic approaches are needed to enhance anticancer effects of available drugs in breast cancer. PMID:26124933

  1. Fenofibrate down-regulates the expressions of androgen receptor (AR) and AR target genes and induces oxidative stress in the prostate cancer cell line LNCaP

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Hu; Zhu, Chen; Qin, Chao [State Key Laboratory of Reproductive Medicine, Department of Urology, First Affiliated Hospital of Nanjing Medical University, Nanjing (China); Tao, Tao [Department of Neurosurgery, First Affiliated Hospital of Nanjing Medical University, Nanjing (China); Li, Jie; Cheng, Gong; Li, Pu; Cao, Qiang; Meng, Xiaoxin; Ju, Xiaobing; Shao, Pengfei; Hua, Lixin [State Key Laboratory of Reproductive Medicine, Department of Urology, First Affiliated Hospital of Nanjing Medical University, Nanjing (China); Gu, Min, E-mail: medzhao1980@163.com [State Key Laboratory of Reproductive Medicine, Department of Urology, First Affiliated Hospital of Nanjing Medical University, Nanjing (China); Yin, Changjun, E-mail: drcjyin@gmail.com [State Key Laboratory of Reproductive Medicine, Department of Urology, First Affiliated Hospital of Nanjing Medical University, Nanjing (China)

    2013-03-08

    Highlights: ► Fenofibrate induces cell cycle arrest in G1 phase and apoptosis in LNCaP cells. ► Fenofibrate reduces the expressions of androgen receptor in LNCaP cells. ► Fenofibrate induces oxidative stress in the prostate cancer cell line LNCaP. -- Abstract: Fenofibrate, a peroxisome proliferator-androgen receptor-alpha agonist, is widely used in treating different forms of hyperlipidemia and hypercholesterolemia. Recent reports have indicated that fenofibrate exerts anti-proliferative and pro-apoptotic properties. This study aims to investigate the effects of fenofibrate on the prostate cancer (PCa) cell line LNCaP. The effects of fenofibrate on LNCaP cells were evaluated by flow cytometry, reverse transcription-polymerase chain reaction, enzyme-linked immunosorbent assays, Western blot analysis, and dual-luciferase reporter assay. Fenofibrate induces cell cycle arrest in G1 phase and apoptosis in LNCaP cells, reduces the expressions of androgen receptor (AR) and AR target genes (prostate-specific antigen and TMPRSS2), and inhibits Akt phosphorylation. Fenofibrate can induce the accumulation of intracellular reactive oxygen species and malondialdehyde, and decrease the activities of total anti-oxidant and superoxide dismutase in LNCaP cells. Fenofibrate exerts an anti-proliferative property by inhibiting the expression of AR and induces apoptosis by causing oxidative stress. Therefore, our data suggest fenofibrate may have beneficial effects in fenofibrate users by preventing prostate cancer growth through inhibition of androgen activation and expression.

  2. Down-regulation of CXCR4 expression by tamoxifen is associated with DNA methyltransferase 3B up-regulation in MCF-7 breast cancer cells.

    Science.gov (United States)

    Kubarek, Ł; Kozłowska, A; Przybylski, M; Lianeri, M; Jagodzinski, P P

    2009-09-01

    The CXCR4 chemokine receptor is a seven transmembrane G protein-coupled receptor present on the surface of various cells including cancer cells. The CXCR4 receptor contributes to the induction of several intracellular signalling pathways that enhance survival, proliferation, and migration of malignant cells. We observed that tamoxifen (Tam) reduced the CXCR4 transcript and protein levels in MCF-7 breast cancer cells. However, we did not see a Tam effect on CXCR4 transcript and protein levels in MCF-7(LVMT3B) cells with RNA interference-mediated knockdown of DNMT3B. We also observed that Tam significantly increased, for several hours, the expression of enzymatically active DNMT3B splice variants in MCF-7 cells. However, there was no Tam effect on these DNMT3B splice variants' expression in MCF-7(LVMT3B) cells. Bisulfite sequencing suggests that Tam may reduce CXCR4 expression via increased methylation of cytosine in the cytosine-guanosine (CpG) dinucleotide island of the CXCR4 promoter of MCF-7 breast cancer cells. Our findings suggest that Tam induces an increase in DNMT3B expression that is associated with the increase of CpG dinucleotide methylation in the CXCR4 promoter and significant reduction of CXCR4 gene expression in MCF-7 cells.

  3. Hyperglycemia Promotes Schwann Cell De-differentiation and De-myelination via Sorbitol Accumulation and Igf1 Protein Down-regulation.

    Science.gov (United States)

    Hao, Wu; Tashiro, Syoichi; Hasegawa, Tomoka; Sato, Yuiko; Kobayashi, Tami; Tando, Toshimi; Katsuyama, Eri; Fujie, Atsuhiro; Watanabe, Ryuichi; Morita, Mayu; Miyamoto, Kana; Morioka, Hideo; Nakamura, Masaya; Matsumoto, Morio; Amizuka, Norio; Toyama, Yoshiaki; Miyamoto, Takeshi

    2015-07-10

    Diabetes mellitus (DM) is frequently accompanied by complications, such as peripheral nerve neuropathy. Schwann cells play a pivotal role in regulating peripheral nerve function and conduction velocity; however, changes in Schwann cell differentiation status in DM are not fully understood. Here, we report that Schwann cells de-differentiate into immature cells under hyperglycemic conditions as a result of sorbitol accumulation and decreased Igf1 expression in those cells. We found that de-differentiated Schwann cells could be re-differentiated in vitro into mature cells by treatment with an aldose reductase inhibitor, to reduce sorbitol levels, or with vitamin D3, to elevate Igf1 expression. In vivo DM models exhibited significantly reduced nerve function and conduction, Schwann cell de-differentiation, peripheral nerve de-myelination, and all conditions were significantly rescued by aldose reductase inhibitor or vitamin D3 administration. These findings reveal mechanisms underlying pathological changes in Schwann cells seen in DM and suggest ways to treat neurological conditions associated with this condition.

  4. Sulforaphene promotes Bax/Bcl2, MAPK-dependent human gastric cancer AGS cells apoptosis and inhibits migration via EGFR, p-ERK1/2 down-regulation.

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    Mondal, Arindam; Biswas, Raktim; Rhee, Yun-Hee; Kim, Jongkee; Ahn, Jin-Chul

    2016-01-01

    Gastric cancer migration and invasion considered as main causes of this cancer-related death around the world. Sulforaphene (4-isothiocyanato-4R-(methylsulfinyl)-1-butene), a structural analog of sulforaphane, has been found to exhibit anticancer potential against different cancers. Our aim was to investigate whether dietary isothiocyanate sulforaphene (SFE) can promote human gastric cancer (AGS) cells apoptosis and inhibit migration. Cells were treated with various concentrations of SFE and cell viability, morphology, intracellular ROS, migration and different signaling protein expressions were investigated. The results indicate that SFE decreases AGS cell viability and induces apoptosis in a dose-dependent manner. Intracellular ROS generation, dose- and time-dependent Bax/Bcl2 alteration and signaling proteins like cytochrome c, Casp-3, Casp-8 and PARP-1 higher expression demonstrated the SFE-induced apoptotic pathway in AGS cells. Again, SFE induced apoptosis also accompanied by the phosphorylation of mitogen-activated protein kinases (MAPKs) like JNK and P-38. Moreover, dose-dependent EGFR, p-ERK1/2 down-regulation and cell migration inhibition at non-toxic concentration confirms SFE activity in AGS cell migration inhibition. Thus, this study demonstrated effective chemotherapeutic potential of SFE by inducing apoptisis as well as inhibiting migration and their preliminary mechanism for human gastric cancer management.

  5. Silencing BMI1 eliminates tumor formation of pediatric glioma CD133+ cells not by affecting known targets but by down-regulating a novel set of core genes.

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    Baxter, Patricia A; Lin, Qi; Mao, Hua; Kogiso, Mari; Zhao, Xiumei; Liu, Zhigang; Huang, Yulun; Voicu, Horatiu; Gurusiddappa, Sivashankarappa; Su, Jack M; Adesina, Adekunle M; Perlaky, Laszlo; Dauser, Robert C; Leung, Hon-chiu Eastwood; Muraszko, Karin M; Heth, Jason A; Fan, Xing; Lau, Ching C; Man, Tsz-Kwong; Chintagumpala, Murali; Li, Xiao-Nan

    2014-01-01

    Clinical outcome of children with malignant glioma remains dismal. Here, we examined the role of over-expressed BMI1, a regulator of stem cell self-renewal, in sustaining tumor formation in pediatric glioma stem cells. Our investigation revealed BMI1 over-expression in 29 of 54 (53.7%) pediatric gliomas, 8 of 8 (100%) patient derived orthotopic xenograft (PDOX) mouse models, and in both CD133+ and CD133- glioma cells. We demonstrated that lentiviral-shRNA mediated silencing of suppressed cell proliferation in vitro in cells derived from 3 independent PDOX models and eliminated tumor-forming capacity of CD133+ and CD133- cells derived from 2 PDOX models in mouse brains. Gene expression profiling showed that most of the molecular targets of BMI1 ablation in CD133+ cells were different from that in CD133- cells. Importantly, we found that silencing BMI1 in CD133+ cells derived from 3 PDOX models did not affect most of the known genes previously associated with the activated BMI1, but modulated a novel set of core genes, including RPS6KA2, ALDH3A2, FMFB, DTL, API5, EIF4G2, KIF5c, LOC650152, C20ORF121, LOC203547, LOC653308, and LOC642489, to mediate the elimination of tumor formation. In summary, we identified the over-expressed BMI1 as a promising therapeutic target for glioma stem cells, and suggest that the signaling pathways associated with activated BMI1 in promoting tumor growth may be different from those induced by silencing BMI1 in blocking tumor formation. These findings highlighted the importance of careful re-analysis of the affected genes following the inhibition of abnormally activated oncogenic pathways to identify determinants that can potentially predict therapeutic efficacy.

  6. Leishmania mexicana promastigotes down regulate JNK and p-38 MAPK activation: Role in the inhibition of camptothecin-induced apoptosis of monocyte-derived dendritic cells.

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    Rodríguez-González, Jorge; Wilkins-Rodríguez, Arturo; Argueta-Donohué, Jesús; Aguirre-García, Magdalena; Gutiérrez-Kobeh, Laila

    2016-04-01

    Dendritic cells (DC) are one of the principal host cells of the obligate intracellular parasite Leishmania. Inhibition of host cell apoptosis is a strategy employed by multiple pathogens to ensure their survival in the infected cell. We have previously shown that the infection of monocyte-derived dendritic cells (moDC) with Leishmania mexicana inhibits campthotecin-induced apoptosis. Nevertheless, the mechanisms involved in the inhibition of apoptosis of dendritic cells by Leishmania have not been established. Mitogen-activated protein kinases (MAPK) are key participants in the process of apoptosis and different species of Leishmania have been shown to regulate these kinases. In the present study, we analyzed the effect of L. mexicana promastigotes in the activation of JNK and p38 MAP kinase and their participation in the inhibition of apoptosis. The infection of moDC with L. mexicana promastigotes diminished significantly the phosphorylation of the MAP kinases JNK and p38. The inhibition of both kinases diminished DNA fragmentation, but in a major extent was the reduction of DNA fragmentation when JNK was inhibited. The capacity of L. mexicana promastigotes to diminish MAP kinases activation is probably one of the strategies employed to delay apoptosis induction in the infected moDC and may have implications for Leishmania pathogenesis by favoring the invasion of its host and the persistence of the parasite in the infected cells.

  7. Down-regulation of ribosomal protein S15A inhibits proliferation of human glioblastoma cells in vivo and in vitro via AKT pathway.

    Science.gov (United States)

    Yao, Yiqun; Liu, Yongjian; Lv, Xiupeng; Dong, Bin; Wang, Feng; Li, Jun; Zhang, Qiuping; Xu, Ruixue; Xu, Yinghui

    2016-04-01

    Ribosomal protein s15a (RPS15A), a highly conserved cytoplasmic protein, promotes mRNA/ribosome interaction in translation. Recent evidence showed that RPS15A is essential for tumor growth. RPS15A expression level was measured in glioblastoma tissue samples and normal brain (NB) tissue samples. RPS15A RNAi stable cell line U87 and U251 was generated by the pLVTHM-GFP lentiviral RNAi expression system. The knockdown efficiency was confirmed by quantitative real-time PCR and western blot. Molecular mechanisms and the effect of RPS15A on cell growth and migration were investigated by using western blot, MTT assay, wound healing assay, transwell migration assay, and tumorigenesis in nude mice. Here, we report that RPS15A is overexpressed in human glioblastoma tumor tissues. RPS15A knockdown inhibits proliferation and migration of glioblastoma cells in vitro. Knocking down RPS15A leads to the level of p-Akt decrease and cell cycle arrested in G0/G1 phase in U87 and U251 cells. Furthermore, the growth of glioblastoma cell-transplanted tumors in nude mice is inhibited by transduction with Lv-shRPS15A. Our findings indicate that RPS15A promotes cell proliferation and migration in glioblastoma for the first time. RPS15A might play a distinct role in glioblastoma and serve as a potential target for therapy.

  8. Comparison of TGFbR2 down-regulation in expanded HSCs on MBA/DBM scaffolds coated by UCB stromal cells.

    Science.gov (United States)

    Hashemi, Zahra Sadat; Moghadam, Mehdi Forouzandeh; Soleimani, Masoud

    2015-05-01

    Bone marrow transplants (BMTs) are mainly limited by a low number of CD34(+) cells. The transforming growth factor-beta (TGF-β) pathway downregulation is a key factor that increases cell self-renewal. In nature, hematopoietic stem cells (HSCs) are in a microenvironment, surrounded by cells in a three-dimensional (3D) configuration. The aim of this study is to investigate the association between a 3D culture and the delivery ratio of downregulation. Demineralized bone matrix (DBM) and mineralized bone allograft (MBA) scaffolds were coated using unrestricted somatic stem cells (USSCs) as the feeder layer. Umbilical cord blood (UCB)-CD34(+) cells were then ex vivo expanded in them and transfected by small interfering RNA (siRNA) against TGFbR2, a type 2 receptor in the TGF-β pathway. Finally, quantitative real-time PCR, flow cytometry, and clonogenic assay were performed. In a global comparison, we observed that the highest expansion ratio, lowest expression level, and the highest CD34 marker belonged to the simple 2D culture transfected group. This suggests that TGFbR2 downregulation in a 2D culture can be done more effectively. The siRNA delivery system and the transfection ratio in an ex vivo environment, which mimicks in vivo conditions, have low efficiency. Genetic modification of the cells needs free 3D spaces to enable better transfection.

  9. Neural cell adhesion molecule-180-mediated homophilic binding induces epidermal growth factor receptor (EGFR) down-regulation and uncouples the inhibitory function of EGFR in neurite outgrowth

    DEFF Research Database (Denmark)

    Povlsen, Gro Klitgaard; Berezin, Vladimir; Bock, Elisabeth

    2008-01-01

    The neural cell adhesion molecule (NCAM) plays important roles in neuronal development, regeneration, and synaptic plasticity. NCAM homophilic binding mediates cell adhesion and induces intracellular signals, in which the fibroblast growth factor receptor plays a prominent role. Recent studies...... not require NCAM-mediated fibroblast growth factor receptor activation....... on axon guidance in Drosophila suggest that NCAM also regulates the epidermal growth factor receptor (EGFR) (Molecular and Cellular Neuroscience, 28, 2005, 141). A possible interaction between NCAM and EGFR in mammalian cells has not been investigated. The present study demonstrates for the first time...

  10. Expression of programmed cell death protein 4 (PDCD4) and miR-21 in urothelial carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Fischer, Nicolas, E-mail: simplissimus@gmx.de [Department of Urology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany); Goeke, Friederike, E-mail: Friederike.goeke@ukb.uni-bonn.de [Department of Pathology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany); Splittstoesser, Vera, E-mail: Veri.sp@web.de [Department of Urology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany); Lankat-Buttgereit, Brigitte, E-mail: Lankatbu@staff.uni-marburg.de [Department of Internal Medicine, Philipps-University of Marburg, Baldingerstrasse, 35043 Marburg (Germany); Mueller, Stefan C., E-mail: Stefan.mueller@ukb.uni-bonn.de [Department of Urology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany); Ellinger, Joerg, E-mail: Joerg.ellinger@ukb.uni-bonn.de [Department of Urology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer The tumor suppressor gene PDCD4 is down-regulated in many tumorous entities. Black-Right-Pointing-Pointer We investigate the impact of PDCD4 and its regulating factor miR-21 in urothelial carcinoma. Black-Right-Pointing-Pointer We confirm PDCD4 as a tumor suppressor gene and it could be a diagnostic marker for this tumor. -- Abstract: Background: We investigated the role of the programmed cell death 4 (PDCD4) tumor suppressor gene in specimens of transitional cell carcinoma and of healthy individuals. Methods: PDCD4 immunohistochemical expression was investigated in 294 cases in histologically proven transitional cell carcinoma in different tumorous stages (28 controls, 122 non-muscle invasive urothelial carcinoma, stages Tis-T1, 119 invasive transitional cell carcinoma stages T2-T4 and 25 metastases). MiR-21 expression, an important PDCD4 regulator, was assessed with real-time PCR analysis and showed inverse correlation to tissue PDCD4 expression. Results: Nuclear and cytoplasmatic PDCD4 immunostaining decreased significantly with histopathological progression of the tumor (p < 0001). Controls showed strong nuclear and cytoplasmatic immunohistochemical staining. MiR-21 up regulation in tissue corresponded to PDCD4 suppression. Conclusions: These data support a decisive role for PDCD4 down regulation in transitional cell carcinoma and confirm miR-21 as a negative regulator for PDCD4. Additionally, PDCD4 immunohistochemical staining turns out to be a possible diagnostic marker for transitional cell carcinoma.

  11. Transcriptomic dissection of tongue squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Schwartz Joel L

    2008-02-01

    Full Text Available Abstract Background The head and neck/oral squamous cell carcinoma (HNOSCC is a diverse group of cancers, which develop from many different anatomic sites and are associated with different risk factors and genetic characteristics. The oral tongue squamous cell carcinoma (OTSCC is one of the most common types of HNOSCC. It is significantly more aggressive than other forms of HNOSCC, in terms of local invasion and spread. In this study, we aim to identify specific transcriptomic signatures that associated with OTSCC. Results Genome-wide transcriptomic profiles were obtained for 53 primary OTSCCs and 22 matching normal tissues. Genes that exhibit statistically significant differences in expression between OTSCCs and normal were identified. These include up-regulated genes (MMP1, MMP10, MMP3, MMP12, PTHLH, INHBA, LAMC2, IL8, KRT17, COL1A2, IFI6, ISG15, PLAU, GREM1, MMP9, IFI44, CXCL1, and down-regulated genes (KRT4, MAL, CRNN, SCEL, CRISP3, SPINK5, CLCA4, ADH1B, P11, TGM3, RHCG, PPP1R3C, CEACAM7, HPGD, CFD, ABCA8, CLU, CYP3A5. The expressional difference of IL8 and MMP9 were further validated by real-time quantitative RT-PCR and immunohistochemistry. The Gene Ontology analysis suggested a number of altered biological processes in OTSCCs, including enhancements in phosphate transport, collagen catabolism, I-kappaB kinase/NF-kappaB signaling cascade, extracellular matrix organization and biogenesis, chemotaxis, as well as suppressions of superoxide release, hydrogen peroxide metabolism, cellular response to hydrogen peroxide, keratinization, and keratinocyte differentiation in OTSCCs. Conclusion In summary, our study provided a transcriptomic signature for OTSCC that may lead to a diagnosis or screen tool and provide the foundation for further functional validation of these specific candidate genes for OTSCC.

  12. Decorin is down-regulated in multiple myeloma and MGUS bone marrow plasma and inhibits HGF-induced myeloma plasma cell viability and migration

    DEFF Research Database (Denmark)

    Kristensen, Ida Bruun; Pedersen, Lise Mariager; Rø, Torstein Baade;

    2013-01-01

    OBJECTIVES: Decorin is a stromal-produced small leucine-rich proteoglycan known to attenuate tumour pro-survival, migration, proliferation and angiogenic signalling pathways. Recent studies have shown that decorin interacts with the hepatocyte growth factor (HGF) receptor c-Met, a potential key p...... of decorin to inhibit HGF-induced effects on MM cell lines were analysed in vitro using cell viability and Transwell migration assays. RESULTS: We found that decorin concentrations were significantly higher (p...

  13. Lack of sik1 in mouse embryonic stem cells impairs cardiomyogenesis by down-regulating the cyclin-dependent kinase inhibitor p57kip2.

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    Antonio Romito

    Full Text Available Sik1 (salt inducible kinase 1 is a serine/threonine kinase that belongs to the stress- and energy-sensing AMP-activated protein kinase family. During murine embryogenesis, sik1 marks the monolayer of future myocardial cells that will populate first the primitive ventricle, and later the primitive atrium suggesting its involvement in cardiac cell differentiation and/or heart development. Despite that observation, the involvement of sik1 in cardiac differentiation is still unknown. We examined the sik1 function during cardiomyocyte differentiation using the ES-derived embryoid bodies. We produced a null embryonic stem cell using a gene-trap cell line carrying an insertion in the sik1 locus. In absence of the sik1 protein, the temporal appearance of cardiomyocytes is delayed. Expression profile analysis revealed sik1 as part of a genetic network that controls the cell cycle, where the cyclin-dependent kinase inhibitor p57(Kip2 is directly involved. Collectively, we provided evidence that sik1-mediated effects are specific for cardiomyogenesis regulating cardiomyoblast cell cycle exit toward terminal differentiation.

  14. Down-regulation of ZnT8 expression in INS-1 rat pancreatic beta cells reduces insulin content and glucose-inducible insulin secretion.

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    Yi Fu

    Full Text Available The SLC30A8 gene codes for a pancreatic beta-cell-expressed zinc transporter, ZnT8. A polymorphism in the SLC30A8 gene is associated with susceptibility to type 2 diabetes, although the molecular mechanism through which this phenotype is manifest is incompletely understood. Such polymorphisms may exert their effect via impacting expression level of the gene product. We used an shRNA-mediated approach to reproducibly downregulate ZnT8 mRNA expression by >90% in the INS-1 pancreatic beta cell line. The ZnT8-downregulated cells exhibited diminished uptake of exogenous zinc, as determined using the zinc-sensitive reporter dye, zinquin. ZnT8-downregulated cells showed reduced insulin content and decreased insulin secretion (expressed as percent of total insulin content in response to hyperglycemic stimulus, as determined by insulin immunoassay. ZnT8-depleted cells also showed fewer dense-core vesicles via electron microscopy. These data indicate that reduced ZnT8 expression in cultured pancreatic beta cells gives rise to a reduced insulin response to hyperglycemia. In addition, although we provide no direct evidence, these data suggest that an SLC30A8 expression-level polymorphism could affect insulin secretion and the glycemic response in vivo.

  15. Squamosamide derivative FLZ inhibits TNF-α-induced ICAM-1 expression via down-regulation of the NF-κB signaling pathway in ARPE-19 cells.

    Science.gov (United States)

    Feng, Ting-Ting; Liang, Ze-Yu; Chen, Song

    2015-01-01

    Dysfunction of the retinal pigment epithelium (RPE) resulting from chronic inflammation is implicated in the pathogenesis of age-related macular degeneration (AMD). It has been reported that tumor necrosis factor-α (TNF-α) could induce intercellular adhesion molecule-1 (ICAM-1) expression in RPE cells. FLZ, a novel synthetic squamosamide derivative from a Chinese herb, Annona glabra, has displayed significant anti-inflammatory activity. However, the effects of FLZ on TNF-α-induced ICAM-1 expression in RPE cells remain unknown. Therefore, in the present study, we evaluated the effects of FLZ on TNF-α-induced ICAM-1 expression in RPE cells. We found that FLZ prevented TNF-α-induced ICAM-1 expression and the ability of monocytes to adhere to ARPE-19 cells induced by TNF-α. Furthermore, FLZ inhibited TNF-α-induced NF-κB p65 expression, as well as phosphorylation of IκBα in ARPE-19 cells. Taken together, these results suggest that FLZ inhibited TNF-α-induced ICAM-1 expression through blocking NF-κB signaling pathway in ARPE-19 cells. Thus, FLZ could be used for designing novel therapeutic agents against AMD.

  16. Trypanosoma cruzi down-regulates lipopolysaccharide-induced MHC class I on human dendritic cells and impairs antigen presentation to specific CD8(+) T lymphocytes.

    Science.gov (United States)

    Van Overtvelt, Laurence; Andrieu, Muriel; Verhasselt, Valérie; Connan, Francine; Choppin, Jeannine; Vercruysse, Vincent; Goldman, Michel; Hosmalin, Anne; Vray, Bernard

    2002-10-01

    Trypanosoma cruzi, the etiological agent of Chagas' disease, may persist for many years in its mammalian host. This suggests escape from the immune response and particularly a suboptimal CD8(+) T cell response, since these cells are involved in infection control. In this report, we show that T. cruzi inhibits the lipopolysaccharide (LPS)-induced up-regulation of MHC class I molecules at the surface of human dendritic cells (DC). To further investigate the functional consequences of this inhibition, a trypomastigote surface antigen-derived peptide (TSA-1(514-522) peptide) was selected for its stable binding to HLA-A*0201 molecules and used to generate a primary T. cruzi-specific human CD8(+) T cell line in vitro. We observed that DC infected with T. cruzi or treated with T. cruzi-conditioned medium (TCM) had a weaker capacity to present this peptide to the specific CD8(+) T cell line as shown in an IFN-gamma ELISPOT assay. Interestingly, T. cruzi or TCM also reduced the antigen presentation capacity of DC to CD8(+) T cell lines specific for the influenza virus M(58-66) or HIV RT(476-484) epitopes. This dysfunction appears to be linked essentially to reduced MHC class I molecule expression since the stimulation of the RT(476-484) peptide-specific CD8(+) T cell line was shown to depend mainly on the MHC class I-TCR interaction and not on the co-stimulatory signals which, however, were also inhibited by T. cruzi. This impairment of DC function may represent a novel mechanism reducing in vivo the host's ability to combat efficiently T. cruzi infection.

  17. Hypoxia-induced down-regulation of microRNA-34a promotes EMT by targeting the Notch signaling pathway in tubular epithelial cells.

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    Rui Du

    Full Text Available BACKGROUND: Hypoxia-induced renal tubular cell epithelial-mesenchymal transition (EMT is an important event leading to renal fibrosis. MicroRNAs (miRNAs are small non-coding RNA molecules that bind to their mRNA targets, thereby leading to translational repression. The role of miRNA in hypoxia-induced EMT is largely unknown. METHODOLOGY/PRINCIPAL FINDINGS: miRNA profiling was performed for the identification of differentially expressed miRNAs in HK-2 cells under normal and low oxygen, and the results were then verified by quantitative real time RT-PCR (qRT-PCR. The function of miRNAs in hypoxia-induced renal tubular cell EMT was assessed by the transfection of specific miRNA inhibitors and mimics. Luciferase reporter gene assays and western blot analysis were performed to validate the target genes of miR-34a. siRNA against Jagged1 was designed to investigate the role of the miR-34a-Notch pathway in hypoxia induced renal tubular cell EMT. miRNA-34a was identified as being downregulated in hypoxic renal tubular epithelial cells. Inhibition of miR-34a expression in HK-2 cells, which highly express endogenous miR-34a, promoted a mesenchymal phenotype accompanied by reduced expression of the epithelial marker Z0-1, E-cadherin and increased expression of the mesenchymal markers α-SMA and vimentin. Conversely, miR-34a mimics effectively prevented hypoxia-induced EMT. Transfection of miRNA-34a in HK-2 cells under hypoxia abolished hypoxia-induced expression of Notch1 and Jagged1 as well as Notch downstream signals, such as snail. Western blot analysis and luciferase reporter gene assays showed direct evidence for miR-34a targeting Notch1 and Jagged1. siRNAs against Jagged1 or Notch1 effectively prevented miR-34a inhibitor-induced tubular epithelial cell EMT. CONCLUSIONS/SIGNIFICANCE: Our study provides evidence that the hypoxia-induced decrease of miR-34a expression could promote EMT in renal tubular epithelial cells by directly targeting Notch1 and

  18. Persistent hepatitis C virus (HCV) infection impairs HCV-specific cytotoxic T cell reactivity through Mcl-1/Bim imbalance due to CD127 down-regulation.

    Science.gov (United States)

    Larrubia, J R; Lokhande, M U; García-Garzón, S; Miquel, J; González-Praetorius, A; Parra-Cid, T; Sanz-de-Villalobos, E

    2013-02-01

    In persistent hepatitis C virus (HCV) infection, HCV-specific cytotoxic T lymphocyte (CTL) reactivity is impaired and this affects HCV control. Interleukin-7 receptor (CD127) expression on these cells could regulate CTL reactivity through Mcl-1/Bim balance modulation. Bim is a pro-apoptotic molecule blocked by the action of Mcl-1. Mcl-1/Bim expression and T cell reactivity on HCV-specific CTLs were compared according to CD127 phenotype. Peripheral blood lymphocytes (PBL) from HLA-A2(+) HCV(+) patients were obtained. HCV-specific CTLs were visualized by staining PBL with anti-CD8 and HLA-A2/peptide pentameric complexes (pentamer). Mcl-1/Bim/CD127 phenotype of HCV-specific CTLs was tested by staining detectable CD8(+)/pentamer(+) cells with anti-Mcl-1/Bim/CD127 antibodies. HCV-specific CTL proliferation ability after specific in vitro challenge was tested in the presence and absence of pancaspase inhibitor z-VAD-fmk. All stained cells were analysed by flow cytometry. CD127(low)-expressing HCV-specific CTLs associated with high HCV viraemia, while CD127(high) correlated with undetectable viral loads (P Bim was up-regulated after antigen encounter (P Bim expression on pentamer(+) cells correlated positively with CD127 expression level (P Bim up-regulation after antigen encounter are involved in CD127(low) HCV-specific CTL hyporeactivity during chronic infection, but it can be overcome by apoptosis blockade.

  19. Galectin-8 Induces Apoptosis in Jurkat T Cells by Phosphatidic Acid-mediated ERK1/2 Activation Supported by Protein Kinase A Down-regulation*

    Science.gov (United States)

    Norambuena, Andrés; Metz, Claudia; Vicuña, Lucas; Silva, Antonia; Pardo, Evelyn; Oyanadel, Claudia; Massardo, Loreto; González, Alfonso; Soza, Andrea

    2009-01-01

    Galectins have been implicated in T cell homeostasis playing complementary pro-apoptotic roles. Here we show that galectin-8 (Gal-8) is a potent pro-apoptotic agent in Jurkat T cells inducing a complex phospholipase D/phosphatidic acid signaling pathway that has not been reported for any galectin before. Gal-8 increases phosphatidic signaling, which enhances the activity of both ERK1/2 and type 4 phosphodiesterases (PDE4), with a subsequent decrease in basal protein kinase A activity. Strikingly, rolipram inhibition of PDE4 decreases ERK1/2 activity. Thus Gal-8-induced PDE4 activation releases a negative influence of cAMP/protein kinase A on ERK1/2. The resulting strong ERK1/2 activation leads to expression of the death factor Fas ligand and caspase-mediated apoptosis. Several conditions that decrease ERK1/2 activity also decrease apoptosis, such as anti-Fas ligand blocking antibodies. In addition, experiments with freshly isolated human peripheral blood mononuclear cells, previously stimulated with anti-CD3 and anti-CD28, show that Gal-8 is pro-apoptotic on activated T cells, most likely on a subpopulation of them. Anti-Gal-8 autoantibodies from patients with systemic lupus erythematosus block the apoptotic effect of Gal-8. These results implicate Gal-8 as a novel T cell suppressive factor, which can be counterbalanced by function-blocking autoantibodies in autoimmunity. PMID:19276072

  20. Bi-phasic effect of interferon (IFN)-alpha: IFN-alpha up- and down-regulates interleukin-4 signaling in human T cells

    DEFF Research Database (Denmark)

    Eriksen, Karsten Wessel; Sommer, Viveca Horst; Woetmann, Anders

    2003-01-01

    Interferon (IFN)-alpha/beta is produced by virally infected cells and is believed to play an important role in early phases of the innate immune response. In addition, IFN-alpha/beta inhibits interleukin (IL)-4 signaling in B cells and monocytes, suggesting that IFN-alpha/beta (like IFN-gamma) is......Interferon (IFN)-alpha/beta is produced by virally infected cells and is believed to play an important role in early phases of the innate immune response. In addition, IFN-alpha/beta inhibits interleukin (IL)-4 signaling in B cells and monocytes, suggesting that IFN-alpha/beta (like IFN......-gamma) is a Th1 cytokine. Here, we study cross-talk between IFN-alpha and IL-4 in human T cells. As expected, stimulation with IFN-alpha for 12-24 h inhibits IL-4 signaling. Surprisingly, however, IFN-alpha has the opposite effect on IL-4 signaling at earlier time points (up to 6 h). Thus, IFN-alpha enhances IL...

  1. Down-regulation of MicroRNAs (MiRs) 203, 887, 3619 and 182 Prevents Vimentin-triggered, Phospholipase D (PLD)-mediated Cancer Cell Invasion.

    Science.gov (United States)

    Fite, Kristen; Gomez-Cambronero, Julian

    2016-01-08

    Breast cancer is a leading cause of morbidity and mortality among women. Metastasis is initiated after epithelial-mesenchymal-transition (EMT). We have found a connection between EMT markers and the expression of four microRNAs (miRs) mediated by the signaling enzyme phospholipase D (PLD). Low aggressive MCF-7 breast cancer cells have low endogenous PLD enzymatic activity and cell invasion, concomitant with high expression of miR-203, -887, and -3619 (that decrease PLD2 translation and a luciferase reporter) and miR-182 (targeting PLD1) that are, therefore, "tumor-suppressor-like" miRs. The combination miR-887+miR-3619 abolished >90% of PLD enzymatic activity. Conversely, post-EMT MDA-MB-231 cells have low miR expression, high levels of PLD1/2, and high aggressiveness. The latter was reversed by ectopically transfecting the miRs, which was negated by silencing miRs with specific siRNAs. We determined that the molecular mechanism is that E-cadherin triggers expression of the miRs in pre-EMT cells, whereas vimentin dampens expression of the miRs in post-EMT invasive cells. This novel work identifies for the first time a set of miRs that are activated by a major pre-EMT marker and deactivated by a post-EMT marker, boosting the transition from low invasion to high invasion, as mediated by the key phospholipid metabolism enzyme PLD.

  2. Leishmania braziliensis-reactive T cells are down-regulated in long-term cured cutaneous Leishmaniasis, but the renewal capacity of T effector memory compartments is preserved.

    Directory of Open Access Journals (Sweden)

    Regina Pereira-Carvalho

    Full Text Available Leishmania (Viannia braziliensis control and tissue damage relate to the effector immune response, which in turn affects clinical outcome. Leishmania reactive CD4(+ and CD8(+ T cells are expanded in long-term healed cutaneous leishmaniasis (hCL patients but their functional characteristics remain to be determined. This study investigates antigen-specific recall in long-term healed CL caused by L. braziliensis infection. Healed CL subjects were grouped according to the time elapsed since the end of therapy: less than two years and two to five years. Activation phenotype (CD69(+ or CD25(+ and subpopulations of memory T cell phenotypes [central memory (Tcm: CD45RO(+ CCR7(+ or effector memory (Tem: CD45RO(+ CCR7(-] were quantified in ex vivo blood mononuclear cells and after Leishmania antigens stimuli. A reduction in the percentage of activated Leishmania-responder CD4(+ and CD8(+ T cells in hCL was associated with the time elapsed since clinical cure. Percentage of CD69(+ in TCD4(+ and TCD8(+ cells were negatively correlated with IL-10 levels. Ex vivo analyses showed contracted Tem CD4(+ and Tem CD8(+ compartments from hCL with long time elapsed since clinical cure, although renewal of these compartments was observed following in vitro exposure to leishmanial stimuli. Our results show that healed L. braziliensis infected patients exhibit a recall response to Leishmania antigens with evident expansion of effector memory T cells. Regulated leishmanial-specific response seems to emerge only about two years after initial contact with the parasite antigens.

  3. The Hereditary Hemochromatosis Protein, HFE, Inhibits Iron Uptake via Down-regulation of Zip14 in HepG2 Cells*

    OpenAIRE

    Gao, Junwei; Zhao, Ningning; Knutson, Mitchell D.; Enns, Caroline A

    2008-01-01

    Lack of functional hereditary hemochromatosis protein, HFE, causes iron overload predominantly in hepatocytes, the major site of HFE expression in the liver. In this study, we investigated the role of HFE in the regulation of both transferrin-bound iron (TBI) and non-transferrin-bound iron (NTBI) uptake in HepG2 cells, a human hepatoma cell line. Expression of HFE decreased both TBI and NTBI uptake. It also resulted in a decrease in the protein levels of Zip14 with no ...

  4. 13-methyltetradecanoic acid exhibits anti-tumor activity on T-cell lymphomas in vitro and in vivo by down-regulating p-AKT and activating caspase-3.

    Directory of Open Access Journals (Sweden)

    Qingqing Cai

    Full Text Available 13-Methyltetradecanoic acid (13-MTD, a saturated branched-chain fatty acid purified from soy fermentation products, induces apoptosis in human cancer cells. We investigated the inhibitory effects and mechanism of action of 13-MTD on T-cell non-Hodgkin's lymphoma (T-NHL cell lines both in vitro and in vivo. Growth inhibition in response to 13-MTD was evaluated by the cell counting kit-8 (CCK-8 assay in three T-NHL cell lines (Jurkat, Hut78, EL4 cells. Flow cytometry analyses were used to monitor the cell cycle and apoptosis. Proteins involved in 13-MTD-induced apoptosis were examined in Jurkat cells by western blotting. We found that 13-MTD inhibited proliferation and induced the apoptosis of T-NHL cell lines. 13-MTD treatment also induced a concentration-dependent arrest of Jurkat cells in the G1-phase. During 13-MTD-induced apoptosis in Jurkat cells, the cleavage of caspase-3 and poly ADP-ribose polymerase (PARP, a caspase enzymolysis product were detected after incubation for 2 h, and increased after extending the incubation time. However, there was no change in the expression of Bcl-2 or c-myc proteins. The appearance of apoptotic Jurkat cells was accompanied by the inhibition of AKT and nuclear factor-kappa B (NF-κB phosphorylation. In addition, 13-MTD could also effectively inhibit the growth of T-NHL tumors in vivo in a xenograft model. The tumor inhibition rate in the experimental group was 40%. These data indicate that 13-MTD inhibits proliferation and induces apoptosis through the down-regulation of AKT phosphorylation followed by caspase activation, which may provide a new approach for treating T-cell lymphomas.

  5. Baicalin protects against TNF-α-induced injury by down-regulating miR-191a that targets the tight junction protein ZO-1 in IEC-6 cells.

    Science.gov (United States)

    Wang, Li; Zhang, Ren; Chen, Jian; Wu, Qihui; Kuang, Zaoyuan

    2017-01-20

    TNF-α plays an important role in the developing process of inflammatory bowel disease. Tight junction protein zonula occludens-1 (ZO-1), one of epithelial junctional proteins, maintains the permeability of intestinal barrier. The objective of this study was to investigate the mechanism of the protective effect of baicalin on TNF-α-induced injury and ZO-1 expression in intestinal epithelial cells (IECs). We found that baicalin pretreatment significantly improved cell viability and cell migration following TNF-α stimulation. MiR-191a inhibitor increased the protective effect of baicalin on cell motility injured by TNF-α. In addition, miR-191a down-regulated the mRNA and protein level of its target gene ZO-1. TNF-α stimulation increased miR-191a expression, leading to the decline of ZO-1 mRNA and protein. Moreover, pretreatment with baicalin reversed TNF-α induced decrease of ZO-1 and increase of miR-191a, miR-191a inhibitor significantly enhanced ZO-1 protein expression restored by baicalin. These results indicate that baicalin exerts a protective effect on IEC-6 cells against TNF-α-induced injury, which is at least partly via inhibiting the expression of miR-191a, thus increasing ZO-1 mRNA and protein levels.

  6. Down-regulation of UDP-glucuronic Acid Biosynthesis Leads to Swollen Plant Cell Walls and Severe Developmental Defects Associated with Changes in Pectic Polysaccharides*

    Science.gov (United States)

    Reboul, Rebecca; Geserick, Claudia; Pabst, Martin; Frey, Beat; Wittmann, Doris; Lütz-Meindl, Ursula; Léonard, Renaud; Tenhaken, Raimund

    2011-01-01

    UDP-glucose dehydrogenase (UGD) plays a key role in the nucleotide sugar biosynthetic pathway, as its product UDP-glucuronic acid is the common precursor for arabinose, xylose, galacturonic acid, and apiose residues found in the cell wall. In this study we characterize an Arabidopsis thaliana double mutant ugd2,3 that lacks two of the four UGD isoforms. This mutant was obtained from a cross of ugd2 and ugd3 single mutants, which do not show phenotypical differences compared with the WT. In contrast, ugd2,3 has a strong dwarfed phenotype and often develops seedlings with severe root defects suggesting that the UGD2 and UGD3 isoforms act in concert. Differences in its cell wall composition in comparison to the WT were determined using biochemical methods indicating a significant reduction in arabinose, xylose, apiose, and galacturonic acid residues. Xyloglucan is less substituted with xylose, and pectins have a reduced amount of arabinan side chains. In particular, the amount of the apiose containing side chains A and B of rhamnogalacturonan II is strongly reduced, resulting in a swollen cell wall. The alternative pathway to UDP-glucuronic acid with the key enzyme myo-inositol oxygenase is not up-regulated in ugd2,3. The pathway also does not complement the ugd2,3 mutation, likely because the supply of myo-inositol is limited. Taken together, the presented data underline the importance of UDP GlcA for plant primary cell wall formation. PMID:21949134

  7. Down-regulation of UDP-glucuronic acid biosynthesis leads to swollen plant cell walls and severe developmental defects associated with changes in pectic polysaccharides.

    Science.gov (United States)

    Reboul, Rebecca; Geserick, Claudia; Pabst, Martin; Frey, Beat; Wittmann, Doris; Lütz-Meindl, Ursula; Léonard, Renaud; Tenhaken, Raimund

    2011-11-18

    UDP-glucose dehydrogenase (UGD) plays a key role in the nucleotide sugar biosynthetic pathway, as its product UDP-glucuronic acid is the common precursor for arabinose, xylose, galacturonic acid, and apiose residues found in the cell wall. In this study we characterize an Arabidopsis thaliana double mutant ugd2,3 that lacks two of the four UGD isoforms. This mutant was obtained from a cross of ugd2 and ugd3 single mutants, which do not show phenotypical differences compared with the WT. In contrast, ugd2,3 has a strong dwarfed phenotype and often develops seedlings with severe root defects suggesting that the UGD2 and UGD3 isoforms act in concert. Differences in its cell wall composition in comparison to the WT were determined using biochemical methods indicating a significant reduction in arabinose, xylose, apiose, and galacturonic acid residues. Xyloglucan is less substituted with xylose, and pectins have a reduced amount of arabinan side chains. In particular, the amount of the apiose containing side chains A and B of rhamnogalacturonan II is strongly reduced, resulting in a swollen cell wall. The alternative pathway to UDP-glucuronic acid with the key enzyme myo-inositol oxygenase is not up-regulated in ugd2,3. The pathway also does not complement the ugd2,3 mutation, likely because the supply of myo-inositol is limited. Taken together, the presented data underline the importance of UDP GlcA for plant primary cell wall formation.

  8. Suppressing Effects of Down-regulating DNMT1 and DNMT3b Expression on the Growth of Human Cholangiocarcinoma Cell Line

    Institute of Scientific and Technical Information of China (English)

    Shi ZUO; Jian LUO; Minfeng LIU; Lining XU; Jingqing DONG; Wei GUO; Shengquan ZOU

    2008-01-01

    Hypermethylation in the promoter region is an important epigenetic mechanism for the transcriptional repression of a number of cancer-associated genes, and over-expression and/or increased activity of DNA methyltransferases are considered to be the main cause of promoter hypermethylation. In order to explore the roles of two methyltransferase members (DNMT1 and DNMT3b) in the cholangiocarcinoma tumorigenesis, antisense eukaryotic expression plasmid of DNMT1 and DNMT3b gene was constructed respectively, and were co-transfected into the human cholangiocarcinoma cell line QBC-939 to observe their biological effects on the cell growth and proliferation ability, apoptosis, cell cycle alteration, and the tumorigenesis ability in the subcutaneous tissue of nude mouse. The results demonstrated that co-transfection with antisense eukaryotic expression plasmid of DNMT1 and DNMT3b gene and single transfection with antisense eukaryotic expression plasmid of DNMT1 gene can suppress the growth and proliferation of QBC-939, block the cell cycle at G1 phase, increase the apoptosis rate, minimize the tumor size in the subcutaneous tissue of nude mouse. The suppressing biological effect of co-transfection is stronger than single transfection with antisense DNMT1. Meanwhile, single transfection with antisense eukaryotic expression plasmid of DNMT3b gene has no effects on the biological characteristics of QBC-939. This study suggests that DNMT1 gene plays a key role in DNA methylation and DNMT3b gene may act as an accessory to support its function in inactivation of tumor suppressor genes. Combination DNMT1 and DNMT3b will inhere their biological effects and have the synergistic effect on suppressing the growth of human cholangiocarcinoma cell line QBC-939.

  9. All Mammalian Hedgehog Proteins Interact with Cell Adhesion Molecule, Down-regulated by Oncogenes (CDO) and Brother of CDO (BOC) in a Conserved Manner*

    OpenAIRE

    Kavran, Jennifer M.; Ward, Matthew D.; Oladosu, Oyindamola O.; Mulepati, Sabin; Leahy, Daniel J.

    2010-01-01

    Hedgehog (Hh) signaling proteins stimulate cell proliferation, differentiation, and tissue patterning at multiple points in animal development. A single Hh homolog is present in Drosophila, but three Hh homologs, Sonic Hh, Indian Hh, and Desert Hh, are present in mammals. Distribution, movement, and reception of Hh signals are tightly regulated, and abnormal Hh signaling is associated with developmental defects and cancer. In addition to the integral membrane proteins Patched and Smoothened, ...

  10. Hydroxy decenoic acid down regulates gtfB and gtfC expression and prevents Streptococcus mutans adherence to the cell surfaces

    Directory of Open Access Journals (Sweden)

    Yousefi Behnam

    2012-07-01

    Full Text Available Abstract Background 10-Hydroxy-2-decenoic acid, an unsaturated fatty acid is the most active and unique component to the royal jelly that has antimicrobial properties. Streptococcus mutans is associated with pathogenesis of oral cavity, gingivoperiodontal diseases and bacteremia following dental manipulations. In the oral cavity, S. mutans colonize the soft tissues including tongue, palate, and buccal mucosa. When considering the role of supragingival dental plaque in caries, the proportion of acid producing bacteria (particularly S. mutans, has direct relevance to the pathogenicity of the plaque. The genes that encode glucosyltransferases (gtfs especially gtfB and gtfC are important in S. mutans colonization and pathogenesis. This study investigated the hydroxy-decenoic acid (HDA effects on gtfB and gtfC expression and S. mutans adherence to cells surfaces. Methods Streptococcus mutans was treated by different concentrations of HPLC purified HDA supplied by Iran Beekeeping and Veterinary Association. Real time RT-PCR and western blot assays were conducted to evaluate gtfB and gtfC genes transcription and translation before and after HDA treatment. The bacterial attachment to the cell surfaces was evaluated microscopically. Results 500 μg ml-1 of HDA inhibited gtfB and gtfC mRNA transcription and its expression. The same concentration of HDA decreased 60% the adherence of S. mutans to the surface of P19 cells. Conclusion Hydroxy-decenoic acid prevents gtfB and gtfC expression efficiently in the bactericide sub-concentrations and it could effectively reduce S. mutans adherence to the cell surfaces. In the future, therapeutic approaches to affecting S. mutans could be selective and it’s not necessary to put down the oral flora completely.

  11. Prostaglandin EP2 receptor signaling protects human trabecular meshwork cells from apoptosis induced by ER stress through down-regulation of p53

    OpenAIRE

    Kalouche, Georges; Boucher, Céline; Coste, Annick; Debussche, Laurent; Orsini, Cécile; Baudouin, Christophe; Debeir, Thomas; Vigé, Xavier; Rostène, William

    2016-01-01

    International audience; E-prostanoid receptor subtype 2 (EP2) agonists are currently under clinical development as hypotensive agents for the treatment of ocular hypertension. However, the effects of EP2 receptor agonists on trabecular meshwork (TM) alterations leading to primary open-angle glaucoma (POAG) are still unknown. Here, we evaluated whether EP2 receptor activation exhibits protective functions on TM cell death induced by endoplasmic reticulum (ER) stress. We show that the EP2 recep...

  12. Selective down-regulation of cell surface cAMP-binding sites and cAMP-induced responses in Dictyostelium discoideum

    NARCIS (Netherlands)

    Kesbeke, Fanja; Haastert, Peter J.M. van

    1985-01-01

    Extracellular cAMP induces an intracellular accumulation of cAMP and cGMP levels in Dictyostelium discoideum, cAMP is detected by cell-surface receptors which are composed of a class of fast-dissociating sites (t1/2 = 1-2 s) and a class of slow-dissociating sites (t1/2 = 15-150 s). Exposure of D. di

  13. A temperature-sensitive allele of a putative mRNA splicing helicase down-regulates many cell wall genes and causes radial swelling in Arabidopsis thaliana.

    Science.gov (United States)

    Howles, Paul A; Gebbie, Leigh K; Collings, David A; Varsani, Arvind; Broad, Ronan C; Ohms, Stephen; Birch, Rosemary J; Cork, Ann H; Arioli, Tony; Williamson, Richard E

    2016-05-01

    The putative RNA helicase encoded by the Arabidopsis gene At1g32490 is a homolog of the yeast splicing RNA helicases Prp2 and Prp22. We isolated a temperature-sensitive allele (rsw12) of the gene in a screen for root radial swelling mutants. Plants containing this allele grown at the restrictive temperature showed weak radial swelling, were stunted with reduced root elongation, and contained reduced levels of cellulose. The role of the protein was further explored by microarray analysis. By using both fold change cutoffs and a weighted gene coexpression network analysis (WGCNA) to investigate coexpression of genes, we found that the radial swelling phenotype was not linked to genes usually associated with primary cell wall biosynthesis. Instead, the mutation has strong effects on expression of secondary cell wall related genes. Many genes potentially associated with secondary walls were present in the most significant WGCNA module, as were genes coding for arabinogalactans and proteins with GPI anchors. The proportion of up-regulated genes that possess introns in rsw12 was above that expected if splicing was unrelated to the activity of the RNA helicase, suggesting that the helicase does indeed play a role in splicing in Arabidopsis. The phenotype may be due to a change in the expression of one or more genes coding for cell wall proteins.

  14. Down-regulation of OsSAG12-1 results in enhanced senescence and pathogen-induced cell death in transgenic rice plants

    Indian Academy of Sciences (India)

    Subaran Singh; Mrunmay Kumar Giri; Praveen Kumar Singh; Adnan Siddiqui; Ashis Kumar Nandi

    2013-09-01

    Senescence is a highly regulated process accompanied by changes in gene expression. While the mRNA levels of most genes decline, the mRNA levels of specific genes (senescence associated genes, SAGs) increase during senescence. Arabidopsis SAG12 (AtSAG12) gene codes for papain-like cysteine protease. The promoter of AtSAG12 is SA-responsive and reported to be useful to delay senescence by expressing cytokinin biosynthesis gene isopentenyltransferase specifically during senescence in several plants including Arabidopsis, lettuce and rice. The physiological role of AtSAG12 is not known; the homozygous atsag12 mutant neither fails to develop senescence-associated vacuoles nor shows any morphological phenotype. Through BLAST search using AtSAG12 amino acid sequences as query, we identified a few putative homologues from rice genome (OsSAGs; Oryza sativa SAGs). OsSAG12-1 is the closest homologue of AtSAG12 with 64% similar amino acid composition. Expression of OsSAG12-1 is induced during senescence and pathogen-induced cell death. To evaluate the possible role of OsSAG12-1 we generated RNAi transgenic lines in Japonica rice cultivar TP309. The transgenic lines developed early senescence at varying levels and showed enhanced cell death when inoculated with bacterial pathogen Xanthomonas oryzae pv.oryzae. Our results suggest that OsSAG12-1 is a negative regulator of cell death in rice.

  15. Down-regulation of ERK1/2 and AKT-mediated X-ray repair cross-complement group 1 protein (XRCC1) expression by Hsp90 inhibition enhances the gefitinib-induced cytotoxicity in human lung cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Tung, Chun-Liang [Department of Pathology, Ditmanson Medical Foundation Chia-Yi Christian Hospital, Chiayi, Taiwan (China); Jian, Yi-Jun [Department of Pathology, Ditmanson Medical Foundation Chia-Yi Christian Hospital, Chiayi, Taiwan (China); Department of Biochemical Science and Technology, National Chiayi University, 300 Syuefu Road, Chiayi 600, Taiwan (China); Syu, Jhan-Jhang; Wang, Tai-Jing; Chang, Po-Yuan; Chen, Chien-Yu; Jian, Yun-Ting [Department of Biochemical Science and Technology, National Chiayi University, 300 Syuefu Road, Chiayi 600, Taiwan (China); Lin, Yun-Wei, E-mail: linyw@mail.ncyu.edu.tw [Department of Biochemical Science and Technology, National Chiayi University, 300 Syuefu Road, Chiayi 600, Taiwan (China)

    2015-05-15

    Gefitinib (Iressa{sup R}, ZD1839) is a selective epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) that blocks growth factor-mediated cell proliferation and extracellular signal-regulated kinases 1/2 (ERK1/2) and AKT signaling activation. It has been shown that inhibition of Hsp90 function can enhance antitumor activity of EGFR-TKI. XRCC1 is an important scaffold protein in base excision repair, which could be regulated by ERK1/2 and AKT pathways. However, the role of ERK1/2 and AKT-mediated XRCC1 expression in gefitinib alone or combination with an Hsp90 inhibitor-induced cytotoxicity in non-small cell lung cancer (NSCLC) cells has not been identified. In this study, gefitinib treatment decreased XRCC1 mRNA and protein expression through ERK1/2 and AKT inactivation in two NSCLC cells, A549 and H1975. Knocking down XRCC1 expression by transfection with small interfering RNA of XRCC1 enhanced the cytotoxicity and cell growth inhibition of gefitinib. Combining treatment of gefitinib with an Hsp90 inhibitor resulted in enhancing the reduction of XRCC1 protein and mRNA levels in gefitinib-exposed A549 and H1975 cells. Compared to a single agent alone, gefitinib combined with an Hsp90 inhibitor resulted in cytotoxicity and cell growth inhibition synergistically in NSCLC cells. Furthermore, transfection with constitutive active MKK1 or AKT vectors rescued the XRCC1 protein level as well as the cell survival suppressed by an Hsp90 inhibitor and gefitinib. These findings suggested that down-regulation of XRCC1 can enhance the sensitivity of gefitinib for NSCLC cells. - Highlights: • Gefitinib treatment decreased XRCC1 mRNA and protein expression in NSCLC cells. • Knocking down XRCC1 expression enhanced the cytotoxic effect of gefitinib. • Gefitinib combined with an Hsp90 inhibitor resulted in synergistically cytotoxicity.

  16. Heat shock protein-27 attenuates foam cell formation and atherogenesis by down-regulating scavenger receptor-A expression via NF-κB signaling.

    Science.gov (United States)

    Raizman, Joshua E; Chen, Yong-Xiang; Seibert, Tara; Hibbert, Benjamin; Cuerrier, Charles M; Salari, Samira; Zhao, Xiaoling; Hu, Tieqiang; Shi, Chunhua; Ma, Xiaoli; Simard, Trevor; Caravaggio, Justin; Rayner, Katey; Bowdish, Dawn; Moore, Kathryn; O'Brien, Edward R

    2013-12-01

    Previously, we showed an inverse correlation between HSP27 serum levels and experimental atherogenesis in ApoE(-/-) mice that over-express HSP27 and speculated that the apparent binding of HSP27 to scavenger receptor-A (SR-A) was of mechanistic importance in attenuating foam cell formation. However, the nature and importance of the interplay between HSP27 and SR-A in atheroprotection remained unclear. Treatment of THP-1 macrophages with recombinant HSP27 (rHSP27) inhibited acLDL binding (-34%; p<0.005) and uptake (-38%, p<0.05). rHSP27 reduced SR-A mRNA (-39%, p=0.02), total protein (-56%, p=0.01) and cell surface (-53%, p<0.001) expression. The reduction in SR-A expression by rHSP27 was associated with a 4-fold increase in nuclear factor-kappa B (NF-κB) signaling (p<0.001 versus control), while an inhibitor of NF-κB signaling, BAY11-7082, attenuated the negative effects of rHSP27 on both SR-A expression and lipid uptake. To determine if SR-A is required for HSP27 mediated atheroprotection in vivo, ApoE(-/-) and ApoE(-/-) SR-A(-/-) mice fed with a high fat diet were treated for 3weeks with rHSP25. Compared to controls, rHSP25 therapy reduced aortic en face and aortic sinus atherosclerotic lesion size in ApoE(-/-) mice by 39% and 36% (p<0.05), respectively, but not in ApoE(-/-)SR-A(-/-) mice. In conclusion, rHSP27 diminishes SR-A expression, resulting in attenuated foam cell formation in vitro. Regulation of SR-A by HSP27 may involve the participation of NF-κB signaling. Lastly, SR-A is required for HSP27-mediated atheroprotection in vivo.

  17. Colostrum whey down-regulates the expression of early and late inflammatory response genes induced by Escherichia coli and Salmonella enterica Typhimurium components in intestinal epithelial cells.

    Science.gov (United States)

    Blais, M; Fortier, M; Pouliot, Y; Gauthier, S F; Boutin, Y; Asselin, C; Lessard, M

    2015-01-28

    Pathogenic invasion by Escherichia coli and Salmonellae remains a constant threat to the integrity of the intestinal epithelium and can rapidly induce inflammatory responses. At birth, colostrum consumption exerts numerous beneficial effects on the properties of intestinal epithelial cells and protects the gastrointestinal tract of newborns from pathogenic invasion. The present study aimed to investigate the effect of colostrum on the early and late inflammatory responses induced by pathogens. The short-term (2 h) and long-term (24 h) effects of exposure to heat-killed (HK) E. coli and Salmonella enterica Typhimurium on gene expression in the porcine intestinal epithelial cell (IPEC-J2) model were first evaluated by microarray and quantitative PCR analyses. Luciferase assays were performed using a NF-κB-luc reporter construct to investigate the effect of colostrum whey treatment on the activation of NF-κB induced by HK bacteria. Luciferase assays were also performed using NF-κB-luc, IL-8-luc and IL-6-luc reporter constructs in human colon adenocarcinoma Caco-2/15 cells exposed to dose-response stimulations with HK bacteria and colostrum whey. Bovine colostrum whey treatment decreased the expression of early and late inflammatory genes induced by HK bacteria in IPEC-J2, as well as the transcriptional activation of NF-κB-luc induced by HK bacteria. Unlike that with colostrum whey, treatment with other milk fractions failed to decrease the activation of NF-κB-luc induced by HK bacteria. Lastly, the reduction of the HK bacteria-induced activation of NF-κB-luc, IL-8-luc and IL-6-luc by colostrum whey was dose dependent. The results of the present study indicate that bovine colostrum may protect and preserve the integrity of the intestinal mucosal barrier in the host by controlling the expression levels of early and late inflammatory genes following invasion by enteric pathogens.

  18. Salvianolic Acid B Down-regulates Matrix Metalloproteinase-9 Activity and Expression in Tumor Necrosis Factor-α-induced Human Coronary Artery Endothelial Cells

    Institute of Scientific and Technical Information of China (English)

    Le Ma; Yun-Qian Guan; Zhong-Dong Du

    2015-01-01

    Background:Salvianolic acid B (Sal B) is a bioactive water-soluble compound of Salviae miltiorrhizae,a traditional herbal medicine that has been used clinically tor the treatment of cardiovascular diseases.This study sought to evaluate the effect of Sal B on matrix metalloproteinase-9 (MMP-9) and on the underlying mechanisms in tumor necrosis factor-α (TNF-α)-activated human coronary artery endothelial cells (HCAECs),a cell model of Kawasaki disease.Methods:HCAECs were pretreated with 1 l0 μmol/L of Sal B,and then stimulated by TNF-α at different time points.The protein expression and activity of MMP-9 were determined by Western blot assay and gelatin zymogram assay,respectively.Nuclear factor-κB (NF-κB) activation was detected with immunofluorescence,electrophoretic mobility shift assay,and Western blot assay.Protein expression levels of mitogen-activated protein kinase (c-Jun N-terminal kinase [JNK],extra-cellular signal-regulated kinase [ERK],and p38) were determined by Western blot assay.Results:After HCAECs were exposed to TNF-α,1-10 μtmol/L Sal B significantly inhibited TNF-α-induced MMP-9 expression and activity.Furthermore,Sal B significantly decreased IκBα phosphorylation and p65 nuclear translocation in HCAECs stimulated with TNF-α for 30 min.In addition,Sal B decreased the phosphorylation of JNK and ERK1/2 proteins in cells treated with TNF-α for 10 min.Conclusions:The data suggested that Sal B suppressed TNF-α-induced MMP-9 expression and activity by blocking the activation of NF-κB,JNK,and ERK1/2 signaling pathways.

  19. Role of TGR-B1-Mediated Down Regulation of NF-kB/Rel Activity During Growth Arrest of Breast Cancer Cells

    Science.gov (United States)

    2001-05-01

    Rogers, P. Toselli , D. Sherr, and G. E. Sonenshein. ROLE OF NF-KB/REL, AHR AND C-MYC IN BREAST CANCER. DOD ERA OF HOPE CONFERENCE 20001, ATLANTA GA...Mo!. Toselli PA, Kim DW, Rogers AE, Sonenshein CE and Cell. Bio!., 4, 1039 1044. Sherr DR. (2000). Breast Cancer Research andl Treatment, Ma Q and...Shneider AM, Toselli P, Lnriiron. Health Persp~ect., 88, 13-- 25. Trombino AF, Yang 5, Hafer Li, Mann KK, Tao XJ, Nebert DW, Petersen DD and PLaga A. (1991

  20. TIMP1 overexpression mediates resistance of MCF-7 human breast cancer cells to fulvestrant and down-regulates progesterone receptor expression

    DEFF Research Database (Denmark)

    Bjerre, Christina; Vinther, Lena; Belling, Kirstine C.

    2013-01-01

    , and cell cycle. Gene and protein expression analyses showed no general defects in estrogen receptor signaling except from lack of progesterone receptor expression and estrogen inducibility in clones with high TIMP1. The present study suggests a relation between high expression level of TIMP1 and loss...... of progesterone receptor expression combined with fulvestrant resistance. Our findings in vitro may have clinical implications as the data suggest that high tumor levels of TIMP1 may be a predictive biomarker for reduced response to fulvestrant....

  1. Grape seed procyanidin reversal of p-glycoprotein associated multi-drug resistance via down-regulation of NF-κB and MAPK/ERK mediated YB-1 activity in A2780/T cells.

    Directory of Open Access Journals (Sweden)

    Bo-xin Zhao

    Full Text Available The expression and function of P-glycoprotein (P-gp is associated with the phenotype of multi-drug resistance (MDR, leading chemotherapy failure of patients suffered with cancer. Grape seed procyanidin(GSP is a natural polyphenol supplement with anti-inflammatory effect. Present study assessed a new use of GSP on the MDR reversal activity and its possible molecular mechanisms in MDR1-overpressing paclitaxel resistant ovarian cancer cells. Our results showed GSP significantly enhanced the cytotoxicity of paclitaxel and adriamycin in paclitaxel resistant A2780/T cells but its parental A2780 cells. Furthermore, GSP strongly inhibited P-gp expression by blocking MDR1 gene transcription, as well as, increased the intracellular accumulation of the P-gp substrate rhodamine-123 in A2780/T cells. Nuclear factor-κB(NF-κB activity, IκB degradation level and NF-κB/p65 nuclear translocation induced by lipopolysaccharide (LPS and receptor activator for nuclear factor-κB ligand (RANKL were markedly inhibited by pre-treatment with GSP. Meanwhile, GSP inhibited MAPK/ERK pathway by decreasing the phosphorylation of ERK1/2, resulting in reduced the Y-box binding protein 1 (YB-1 activation with blocking its nuclear translocation. Moreover, the up-regulation of P-gp expression, the activation of AKT/NF-κB and MAPK/ERK pathway induced by LPS was attenuated by GSP administration. Compared with PDTC and U1026, inhibitor of NF-κB and MAPK/ERK respectively, GSP showed the same tendency of down-regulating NF-κB and MAPK/ERK mediated YB-1 activities. Thus, GSP reverses P-gp associated MDR by inhibiting the function and expression of P-gp through down-regulation of NF-κB activity and MAPK/ERK pathway mediated YB-1 nuclear translocation, offering insight into the mechanism of reversing MDR by natural polyphenol supplement compounds. GSP could be a new potential MDR reversal agent used for combination therapy with chemotherapeutics in clinic.

  2. Resveratrol Inhibits the Invasion of Glioblastoma-Initiating Cells via Down-Regulation of the PI3K/Akt/NF-κB Signaling Pathway

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    Yuming Jiao

    2015-06-01

    Full Text Available Invasion and metastasis of glioblastoma-initiating cells (GICs are thought to be responsible for the progression and recurrence of glioblastoma multiforme (GBM. A safe drug that can be applied during the rest period of temozolomide (TMZ maintenance cycles would greatly improve the prognosis of GBM patients by inhibiting GIC invasion. Resveratrol (RES is a natural compound that exhibits anti-invasion properties in multiple tumor cell lines. The current study aimed to evaluate whether RES can inhibit GIC invasion in vitro and in vivo. GICs were identified using CD133 and Nestin immunofluorescence staining and tumorigenesis in non-obese diabetic severe combined immunodeficient (NOD/SCID mice. Invasive behaviors, including the adhesion, invasion and migration of GICs, were determined by tumor invasive assays in vitro and in vivo. The activity of matrix metalloproteinases (MMPs was measured by the gelatin zymography assay. Western blotting analysis and immunofluorescence staining were used to determine the expression of signaling effectors in GICs. We demonstrated that RES suppressed the adhesion, invasion and migration of GICs in vitro and in vivo. Moreover, we proved that RES inhibited the invasion of GICs via the inhibition of PI3K/Akt/NF-κB signal transduction and the subsequent suppression of MMP-2 expression.

  3. Naringenin modulates the metastasis of human prostate cancer cells by down regulating the matrix metalloproteinases -2/-9 via ROS/ERK1/2 pathways

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    Er-Jiang Lin

    2014-08-01

    Full Text Available Metastasis is a multifactorial condition that complicates cancer treatment options and widens the target of treatment. Matrix mettalopriteinases (MMPs of the extracellular matrix (ECM are involved in metastasis, thus they present as potential targets in halting cancer metastasis. The study was undertaken to investigate the influence of naringenin, a naturally occurring flavonoid on the metastasis of human prostate cancer cells (PC-3 and DU145. Naringenin was observed to be effective in reducing the viability and migratory percentage of PC-3 and DU145 cells. Naringenin significantly reduced the expression and activities of the chief MMPs (MMP-2 and MMP-9 as assessed by western blotting, real-time PCR and gelatin zymography analysis. The influence of naringenin on extracellular signal-regulated kinase (ERK -ERK1/2 was analysed by western blotting. The results indicated that naringenin was able to effectively inhibit ERK1/2. Naringenin exposure also significantly suppressed the levels of reactive oxygen species (ROS. Naringenin thus stands as an effective chemotherapeutic agent for prostate cancer treatment that could be further explored.

  4. Magnolol inhibits colonic motility through down-regulation of voltage-sensitive L-type Ca2+ channels of colonic smooth muscle cells in rats.

    Science.gov (United States)

    Zhang, Man; Zang, Kai-Hong; Luo, Jia-Lie; Leung, Fung-Ping; Huang, Yu; Lin, Cheng-Yuan; Yang, Zhi-Jun; Lu, Ai-Ping; Tang, Xu-Dong; Xu, Hong-Xi; Sung, Joseph Jao-yiu; Bian, Zhao-Xiang

    2013-11-15

    This study aimed to investigate the effect of magnolol (5,5'-diallyl-2,2'-biphenyldiol) on contraction in distal colonic segments of rats and the underlying mechanisms. Colonic segments were mounted in organ baths for isometric force measurement. Whole-cell voltage-sensitive L-type Ca(2+) currents were recorded on isolated single colonic smooth muscle cells using patch-clamp technique. The spontaneous contractions and acetylcholine (ACh)- and Bay K 8644-induced contractions were inhibited by magnolol (3-100 μM). In the presence of Bay K8644 (100 nM), magnolol (10-100 μM) inhibited the contraction induced by 10 μM ACh. By contrast, tetrodotoxin (100 nM) and Nώ-nitro-L-arginine methyl ester (L-NAME 100 μM) did not change the inhibitory effect of magnolol (10 μM). In addition, magnolol (3-100 μM) inhibited the L-type Ca(2+) currents. The present results suggest that magnolol inhibits colonic smooth muscle contraction through downregulating L-type Ca(2+) channel activity.

  5. Down-regulated NOD2 by immunosuppressants in peripheral blood cells in patients with SLE reduces the muramyl dipeptide-induced IL-10 production.

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    Shui-Lian Yu

    Full Text Available BACKGROUND: Pattern recognition receptors (PRRs such as Toll-like receptors are aberrantly expressed of peripheral blood mononuclear cells (PBMCs in systemic lupus erythematosus (SLE patients, for playing immunopathological roles. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the expression and function of the PRR nucleotide-binding oligomerization domain (NOD2 in SLE. NOD2 expression in T, B lymphocytes, monocytes, myeloid dendritic cells (mDCs and plasmacytoid dendritic cells (pDCs was assessed in SLE patients and healthy controls (HCs using flow cytometric analysis. Ex vivo production of cytokines from PBMCs upon NOD2 agonist muramyl dipeptide (MDP stimulation was assessed using Cytometric Bead Array. Over-expression of NOD2 in monocytes was observed in immunosuppressant naïve SLE patients, and was positively associated with longer disease duration. Immunosuppressive therapy was an independent explanatory variable for downregulating NOD2 expression in CD8+ T, monocytes, mDCs and pDCs. Ex vivo basal productions of cytokines (IL-6, IL-8 and IL-10 were significantly increased in immunosuppressant naïve patients and patients with active disease despite immunosuppressants compared with HCs. Upon MDP stimulaiton, relative induction (% of cytokines (IL-1β from PBMC was significantly increased in immunosuppressant naïve patients with inactive disease, and patients with active disease despite immunosuppressant treatment compared with HCs. Immunosuppressant usage was associated with a decreased basal production and MDP induced relative induction (% of IL-10 in patients with inactive disease compared with immunosuppressant naïve patients and HCs. CONCLUSIONS/SIGNIFICANCE: Bacterial exposure may increase the NOD2 expression in monocytes in immunosuppressant naïve SLE patients which can subsequently lead to aberrant activation of PBMCs to produce proinflammatory cytokines, implicating the innate immune response for extracellular pathogens in the

  6. BME, a novel compound of anthraquinone, down regulated P-glycoprotein expression in doxorubicin-resistant human myelogenous leukemia (K562/DOX) cells via generation of reactive oxygen species.

    Science.gov (United States)

    Wang, Jianhong; Liu, Lu; Cen, Juan; Ji, Biansheng

    2015-09-01

    P-glycoprotein (P-gp)-mediated multidrug resistance (MDR) in tumor cells is still a main obstacle for the chemotherapeutic treatment of cancers. Thus, development of effective MDR reversing agents is an important approach in the clinic. The present study revealed that BME, a novel compound of anthraquinone, elevated intracellular accumulation of the P-gp substrates and reduced concentration resulting in 50% inhibition of cell growth (IC50) values for doxorubicin (DOX) in doxorubicin-resistant human myelogenous leukemia (K562/DOX) cells. Further more, BME was also reported to down regulated P-gp expression accompanying with generation of nontoxic low level of intracellular reactive oxygen species (iROS) and activation of extracellular signal-regulated kinase (ERK)1/2 as well as c-JUN N-terminal kinase (JNK). However, treatment with N-acetyl-cysteine (NAC), U0216 and SP600125 almost abolished actions of the BME mentioned above. These results indicated that the effect of the BME on the P-gp may be involved in generation of nontoxic low level of iROS and activation of ERK1/2 or JNK, which suggested valuable clues to screen and develop P-gp reversing agents.

  7. MiR-138 promotes smooth muscle cells proliferation and migration in db/db mice through down-regulation of SIRT1

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    Xu, Juan [Department of Gynecology, Changzhou Maternity and Children Health Hospital, Changzhou, Jiangsu 213003 (China); Li, Li; Yun, Hui-fang [Department of Anesthesiology, Changzhou No. 2 People' s Hospital, Changzhou, Jiangsu 213003 (China); Han, Ye-shan, E-mail: yeshanhan123@163.com [Department of Anesthesiology, Changzhou No. 2 People' s Hospital, Changzhou, Jiangsu 213003 (China)

    2015-08-07

    Background: Diabetic vascular smooth muscle cells (VSMCs) exhibit significantly increased rates of proliferation and migration, which was the most common pathological change in atherosclerosis. In addition, the study about the role for miRNAs in the regulation of VSMC proliferation is just beginning to emerge and additional miRNAs involved in VSMC proliferation modulation should be identified. Methods: The expression of miR-138 and SIRT1 were examined in SMCs separated from db/db mice and in SMC lines C-12511 exposed to high glucose with qRT-PCR and western blot. The regulation of miR-138 on the expression of SMCs was detected with luciferase report assay. VSMCs proliferation and migration assays were performed to examine the effect of miR-138 inhibitor on VSMCs proliferation and migration. Results: We discovered that higher mRNA level of miR-138 and reduced expression of SIRT1 were observed in SMCs separated from db/db mice and in SMC lines C-12511. Moreover, luciferase report assay showed that the activity of SIRT1 3′-UTR was highly increased by miR-138 inhibitor and reduced by miR-138 mimic. In addition, we examined that the up-regulation of NF-κB induced by high glucose in SMCs was reversed by resveratrol and miR-138 inhibitor. MTT and migration assays showed that miR-138 inhibitor attenuated the proliferation and migration of smooth muscle cells. Conclusion: In this study, we revealed that miR-138 might promote proliferation and migration of SMC in db/db mice through suppressing the expression of SIRT1. - Highlights: • Higher mRNA level of miR-138 was observed in SMCs from db/db mice. • The mRNA and protein level of SIRT1 in SMCs from db/db mice were greatly reduced. • miR-138 could regulate the expression of SIRT1 in SMCs. • SIRT1 overexpression reversed the up-regulation of acetylized p65 and NF-κB induced by high glucose. • MiR-138 inhibitor reversed VSMCs proliferation and migration induced by high glucose.

  8. Beta-elemene blocks epithelial-mesenchymal transition in human breast cancer cell line MCF-7 through Smad3-mediated down-regulation of nuclear transcription factors.

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    Xian Zhang

    Full Text Available Epithelial-mesenchymal transition (EMT is the first step required for breast cancer to initiate metastasis. However, the potential of drugs to block and reverse the EMT process are not well explored. In the present study, we investigated the inhibitory effect of beta-elemene (ELE, an active component of a natural plant-derived anti-neoplastic agent in an established EMT model mediated by transforming growth factor-beta1 (TGF-β1. We found that ELE (40 µg/ml blocked the TGF-β1-induced phenotypic transition in the human breast cancer cell line MCF-7. ELE was able to inhibit TGF-β1-mediated upregulation of mRNA and protein expression of nuclear transcription factors (SNAI1, SNAI2, TWIST and SIP1, potentially through decreasing the expression and phosphorylation of Smad3, a central protein mediating the TGF-β1 signalling pathway. These findings suggest a potential therapeutic benefit of ELE in treating basal-like breast cancer.

  9. Down-regulation of the Nucleotide Excision Repair gene XPG as a new mechanism of drug resistance in human and murine cancer cells

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    Geroni Cristina

    2010-09-01

    Full Text Available Abstract Background Drug resistance is one of the major obstacles limiting the activity of anticancer agents. Activation of DNA repair mechanism often accounts for increase resistance to cancer chemotherapy. Results We present evidence that nemorubicin, a doxorubicin derivative currently in clinical evaluation, acts through a mechanism of action different from classical anthracyclines, requiring an intact nucleotide excision repair (NER system to exert its activity. Cells made resistant to nemorubicin show increased sensitivity to UV damage. We have analysed the mechanism of resistance and discovered a previously unknown mechanism resulting from methylation-dependent silencing of the XPG gene. Restoration of NER activity through XPG gene transfer or treatment with demethylating agents restored sensitivity to nemorubicin. Furthermore, we found that a significant proportion of ovarian tumors present methylation of the XPG promoter. Conclusions Methylation of a NER gene, as described here, is a completely new mechanism of drug resistance and this is the first evidence that XPG gene expression can be influenced by an epigenetic mechanism. The reported methylation of XPG gene could be an important determinant of the response to platinum based therapy. In addition, the mechanism of resistance reported opens up the possibility of reverting the resistant phenotype using combinations with demethylating agents, molecules already employed in the clinical setting.

  10. Resveratrol and its methoxy derivatives modulate the expression of estrogen metabolism enzymes in breast epithelial cells by AhR down-regulation.

    Science.gov (United States)

    Licznerska, Barbara; Szaefer, Hanna; Wierzchowski, Marcin; Sobierajska, Hanna; Baer-Dubowska, Wanda

    2017-01-01

    Our earlier studies have shown that compared to resveratrol, its analogs with ortho-methoxy substituents exert stronger antiproliferative and proapoptotic activity. Since estrogens are considered the major risk factors of breast carcinogenesis, the aim of this study was to evaluate the effect of 3,4,2'-trimethoxy (3MS), 3,4,2',4'-tetramethoxy (4MS), and 3,4,2',4',6'-pentamethoxy (5MS) trans-stilbenes on the constitutive expression of the enzymes involved in estrogen metabolism, as well as receptors: AhR and HER2 in breast epithelial cell line MCF10A. The results showed different effect of resveratrol and its methoxy derivatives on the expression of genes encoding key enzymes of estrogen synthesis and catabolism. Resveratrol at the doses of 1 and 5 µmol/L increased the level of CYP19 transcript and protein level, while 5MS reduced mRNA transcript of both CYP19 and STS genes. Resveratrol and all its derivatives reduced also SULT1E1 mRNA transcript level. The reduced expression of AhR, CYP1A1, and 1B1 was also found as a result of treatment with these compounds. The most significant changes were found in the case of AhR. The most potent inhibitor of CYP1A1 and 1B1 genes expression was 5MS, which reduced the levels of mRNA transcript and protein of both CYPs from 31 to 89% of the initial levels. These results indicate that methoxy derivatives of resveratrol might be efficient modulators of estrogen metabolism. Moreover, the number of methoxy groups introduced to stilbene structure may play a certain role in this effect.

  11. Phorbol Esters from Jatropha Meal Triggered Apoptosis, Activated PKC-δ, Caspase-3 Proteins and Down-Regulated the Proto-Oncogenes in MCF-7 and HeLa Cancer Cell Lines

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    Syahida Ahmad

    2012-09-01

    Full Text Available Jatropha meal produced from the kernel of Jatropha curcas Linn. grown in Malaysia contains phorbol esters (PEs. The potential benefits of PEs present in the meal as anticancer agent are still not well understood. Hence, this study was conducted to evaluate the cytotoxic effects and mode of actions of PEs isolated from Jatropha meal against breast (MCF-7 and cervical (HeLa cancer cell lines. Isolated PEs inhibited cells proliferation in a dose-dependent manner of both MCF-7 and HeLa cell lines with the IC50 of 128.6 ± 2.51 and 133.0 ± 1.96 µg PMA equivalents/mL respectively, while the values for the phorbol 12-myristate 13-acetate (PMA as positive control were 114.7 ± 1.73 and 119.6 ± 3.73 µg/mL, respectively. Microscopic examination showed significant morphological changes that resemble apoptosis in both cell lines when treated with PEs and PMA at IC50 concentration after 24 h. Flow cytometry analysis and DNA fragmentation results confirmed the apoptosis induction of PEs and PMA in both cell lines. The PEs isolated from Jatropha meal activated the PKC-δ and down-regulated the proto-oncogenes (c-Myc, c-Fos and c-Jun. These changes probably led to the activation of Caspase-3 protein and apoptosis cell death occurred in MCF-7 and HeLa cell lines upon 24 h treatment with PEs and PMA. Phorbol esters of Jatropha meal were found to be promising as an alternative to replace the chemotherapeutic drugs for cancer therapy.

  12. Phorbol esters from Jatropha meal triggered apoptosis, activated PKC-δ, caspase-3 proteins and down-regulated the proto-oncogenes in MCF-7 and HeLa cancer cell lines.

    Science.gov (United States)

    Oskoueian, Ehsan; Abdullah, Norhani; Ahmad, Syahida

    2012-09-10

    Jatropha meal produced from the kernel of Jatropha curcas Linn. grown in Malaysia contains phorbol esters (PEs). The potential benefits of PEs present in the meal as anticancer agent are still not well understood. Hence, this study was conducted to evaluate the cytotoxic effects and mode of actions of PEs isolated from Jatropha meal against breast (MCF-7) and cervical (HeLa) cancer cell lines. Isolated PEs inhibited cells proliferation in a dose-dependent manner of both MCF-7 and HeLa cell lines with the IC₅₀ of 128.6 ± 2.51 and 133.0 ± 1.96 µg PMA equivalents/mL respectively, while the values for the phorbol 12-myristate 13-acetate (PMA) as positive control were 114.7 ± 1.73 and 119.6 ± 3.73 µg/mL, respectively. Microscopic examination showed significant morphological changes that resemble apoptosis in both cell lines when treated with PEs and PMA at IC₅₀ concentration after 24 h. Flow cytometry analysis and DNA fragmentation results confirmed the apoptosis induction of PEs and PMA in both cell lines. The PEs isolated from Jatropha meal activated the PKC-δ and down-regulated the proto-oncogenes (c-Myc, c-Fos and c-Jun). These changes probably led to the activation of Caspase-3 protein and apoptosis cell death occurred in MCF-7 and HeLa cell lines upon 24 h treatment with PEs and PMA. Phorbol esters of Jatropha meal were found to be promising as an alternative to replace the chemotherapeutic drugs for cancer therapy.

  13. Synchronous thyroid carcinoma and squamous cell carcinoma. A case report

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jae Seo [Chonnam National Univ. School of Dentistry, Kwangju (Korea, Republic of)

    2006-12-15

    Thyroid carcinoma occurring as a second primary associated with head and neck squamous cell carcinoma (SCC) is unusual. This report presents a synchronous thyroid carcinoma and squamous cell carcinoma in the anterior palate region of a 41-year-old man. The clinical, radiologic, and histologic features are described. At 10-month follow-up after operation, no evidence of recurrence ana metastasis was present.

  14. CMTM5 exhibits tumor suppressor activity through promoter methylation in oral squamous cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Heyu [Central Laboratory, Peking University School of Stomatology, Beijing (China); Nan, Xu [Center for Human Disease Genomics, Department of Immunology, Key Laboratory of Medical Immunology, Ministry of Health, School of Basic Medical Sciences, Peking University, Beijing (China); Li, Xuefen [Central Laboratory, Peking University School of Stomatology, Beijing (China); Chen, Yan; Zhang, Jianyun [Department of Oral Pathology, Peking University School of Stomatology, Beijing (China); Sun, Lisha [Central Laboratory, Peking University School of Stomatology, Beijing (China); Han, Wenlin [Center for Human Disease Genomics, Department of Immunology, Key Laboratory of Medical Immunology, Ministry of Health, School of Basic Medical Sciences, Peking University, Beijing (China); Li, Tiejun, E-mail: litiejun22@vip.sina.com [Department of Oral Pathology, Peking University School of Stomatology, Beijing (China)

    2014-05-02

    Highlights: • Down-regulation of CMTM5 expression in OSCC tissues was found. • The promoter methylation status of CMTM5 was measured. • CMTM5-v1 inhibited cell proliferation and migration and induced apoptosis. • CMTM5 might act as a putative tumor suppressor gene in OSCC. - Abstract: Oral squamous cell carcinoma (OSCC) is one of the most common types of malignancies in the head and neck region. CKLF-like MARVEL transmembrane domain-containing member 5 (CMTM5) has been recently implicated as a tumor suppressor gene in several cancer types. Herein, we examined the expression and function of CMTM5 in oral squamous cell carcinoma. CMTM5 was down-regulated in oral squamous cell lines and tumor samples from patients with promoter methylation. Treatment with the demethylating agent 5-aza-2′-deoxycytidine restored CMTM5 expression. In the OSCC cell lines CAL27 and GNM, the ectopic expression of CMTM5-v1 strongly inhibited cell proliferation and migration and induced apoptosis. In addition, CMTM5-v1 inhibited tumor formation in vivo. Therefore, CMTM5 might act as a putative tumor suppressor gene through promoter methylation in oral squamous cell carcinoma.

  15. MiR-193a-3p and miR-193a-5p suppress the metastasis of human osteosarcoma cells by down-regulating Rab27B and SRR, respectively.

    Science.gov (United States)

    Pu, Youguang; Zhao, Fangfang; Cai, Wenjing; Meng, Xianghui; Li, Yinpeng; Cai, Shanbao

    2016-04-01

    MicroRNAs have been identified as key players in the development and progression of osteosarcoma, which is the most common primary malignancy of bone. Sequencing-based miR-omic and quantitative real-time PCR analyses suggested that the expression of miR-193a-3p and miR-193a-5p was decreased by DNA methylation at their promoter region in a highly metastatic osteosarcoma cell line (MG63.2) relative to their expression in the less metastatic MG63 cell line. Further wound-healing and invasion assays demonstrated that both miR-193a-3p and miR-193a-5p suppressed osteosarcoma cell migration and invasion. Moreover, introducing miR-193a-3p and miR-193a-5p mimics into MG63.2 cells or antagomiRs into MG63 cells confirmed their critical roles in osteosarcoma metastasis. Additionally, bioinformatics prediction along with biochemical assay results clearly suggested that the secretory small GTPase Rab27B and serine racemase (SRR) were direct targets of miR-193a-3p and miR-193a-5p, respectively. These two targets are indeed involved in the miR-193a-3p- and miR-193a-5p-induced suppression of osteosarcoma cell migration and invasion. MiR-193a-3p and miR-193a-5p play important roles in osteosarcoma metastasis through down-regulation of the Rab27B and SRR genes and therefore may serve as useful biomarkers for the diagnosis of osteosarcoma and as potential candidates for the treatment of metastatic osteosarcoma.

  16. 松果菊苷通过下调p53的表达抑制人成纤维细胞的衰老%Echinacoside suppresses cellular senescence of human fibroblastic cells by down-regulation of p53

    Institute of Scientific and Technical Information of China (English)

    朱慧; 成聪; 张弛; 王钊

    2011-01-01

    松果菊苷是一种从肉苁蓉中分离得到的苯乙醇苷类化合物,前期研究结果表明其有延缓人成纤维细胞衰老的作用.为了阐明松果菊苷抗衰老的机制,我们对相关基因p53,p16,p21及Rb的mRNA及蛋白水平的表达进行了检测,结果表明用松果菊苷处理后,衰老的人成纤维细胞(MRC-5) p53的表达被下调,且呈剂量依赖方式.进一步的研究显示可能和SIRTI蛋白的上调有关.分子对接模拟结果显示松果菊苷的作用可能优于另一公认的抗衰老剂白藜芦醇.松果菊苷可能是一种潜在的可以调控细胞衰老的化合物.%Echinacoside is one of the phenylethanoids isolated from the stems of Cistanches salsa.Our previous research showed that echinacoside has anti-senescence activity.To investigate the mechanism of echinacoside's anti-senescence activity,the expressions of p53,p21,p16 and Rb at the mRNA and protein levels were determined.Results showed that the expression of p53 was down-regulated significantly in a dose dependent manner after treatment with echinacoside.Further experiments suggested that the down-regulation of p53 may be correlated with the uprcgulation of SIRT1.In addition,echinacoside may exhibit considerable higher affinity towards SIRT1 than resveratrol,according to our molecular docking simulation.In conclusion,we expect that echinacoside might be a promising candidate for regulating cell senescence.

  17. Down-regulation of 14-3-3β exerts anti-cancer effects through inducing ER stress in human glioma U87 cells: Involvement of CHOP–Wnt pathway

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    Cao, Lei; Lei, Hui; Chang, Ming-Ze; Liu, Zhi-Qin [Department of Neurological Disease, Xi' an Central Hospital, Xi' an Jiaotong University, Xi' an, Shaanxi 710000 (China); Bie, Xiao-Hua, E-mail: biexiaohua_xjtu@126.com [Department of Functional Neurosurgery, Xi' an Red Cross Hospital, Xi' an Jiaotong University, Xi' an, Shaanxi 710054 (China)

    2015-07-10

    We previously identified 14-3-3β as a tumor-specific isoform of 14-3-3 protein in astrocytoma, but its functional role in glioma cells and underlying mechanisms are poorly understood. In the present study, we investigated the effects of 14-3-3β inhibition in human glioma U87 cells using specific targeted small interfering RNA (siRNA). The results showed that 14-3-3β is highly expressed in U87 cells but not in normal astrocyte SVGp12 cells. Knockdown of 14-3-3β by Si-14-3-3β transfection significantly decreased the cell viability but increased the LDH release in a time-dependent fashion in U87 cells, and these effects were accompanied with G0/G1 cell cycle arrest and apoptosis. In addition, 14-3-3β knockdown induced ER stress in U87 cells, as evidenced by ER calcium release, increased expression of XBP1S mRNA and induction of ER related pro-apoptotic factors. Down-regulation of 14-3-3β significantly decreased the nuclear localization of β-catenin and inhibited Topflash activity, which was shown to be reversely correlated with CHOP. Furthermore, Si-CHOP and sFRP were used to inhibit CHOP and Wnt, respectively. The results showed that the anti-cancer effects of 14-3-3β knockdown in U87 cells were mediated by increased expression of CHOP and followed inhibition of Wnt/β-catenin pathway. In summary, the remarkable efficiency of 14-3-3β knockdown to induce apoptotic cell death in U87 cells may find therapeutic application for the treatment of glioma patients. - Highlights: • Knockdown of 14-3-3β leads to cytotoxicity in human glioma U87 cells. • Knockdown of 14-3-3β induces cell cycle arrest and apoptosis in U87 cells. • Knockdown of 14-3-3β results in ER stress in U87 cells. • Knockdown of 14-3-3β inhibits Wnt/β-catenin pathway via CHOP activation.

  18. Down-regulation of beta1,4-galactosyltransferase V is a critical part of etoposide-induced apoptotic process and could be mediated by decreasing Sp1 levels in human glioma cells.

    Science.gov (United States)

    Jiang, Jianhai; Shen, Jialin; Wu, Tao; Wei, Yuanyan; Chen, Xiaoning; Zong, Hongliang; Zhang, Si; Sun, Maoyun; Xie, Jianhui; Kong, Xiangfei; Yang, Yanzhong; Shen, Aiguo; Wang, Hanzhou; Gu, Jianxin

    2006-11-01

    beta1,4-Galactosyltransferase V (beta1,4GalT V; EC 2.4.1.38) is considered to be very important in glioma for expressing transformation-related highly branched N-glycans. Recently, we have characterized beta1,4GalT V as a positive growth regulator in several glioma cell lines. However, the role of beta1,4GalT V in glioma therapy has not been clearly reported. In this study, interfering with the expression of beta1,4GalT V by its antisense cDNA in SHG44 human glioma cells markedly promoted apoptosis induced by etoposide and the activation of caspases as well as processing of Bid and expression of Bax and Bak. Conversely, the ectopic expression of beta1,4GalT V attenuated the apoptotic effect of etoposide on SHG44 cells. In addition, both the beta1,4GalT V transcription and the binding of total or membrane glycoprotein with Ricinus communis agglutinin-I (RCA-I) were partially reduced in etoposide-treated SHG44 cells, correlated well with a decreased level of Sp1 that has been identified as an activator of beta1,4GalT V transcription. Collectively, our results suggest that the down-regulation of beta1,4GalT V expression plays an important role in etoposide-induced apoptosis and could be mediated by a decreasing level of Sp1 in SHG44 cells, indicating that inhibitors of beta1,4GalT V may enhance the therapeutic efficiency of etoposide for malignant glioma.

  19. Exposure of E. coli to DNA-methylating agents impairs biofilm formation and invasion of eukaryotic cells via down regulation of the N-acetylneuraminate lyase NanA

    Directory of Open Access Journals (Sweden)

    Pamela eDi Pasquale

    2016-02-01

    Full Text Available DNA methylation damage can be induced by endogenous and exogenous chemical agents, which has led every living organism to develop suitable response strategies. We investigated protein expression profiles of Escherichia coli upon exposure to the alkylating agent methyl-methane sulfonate (MMS by differential proteomics. Quantitative proteomic data showed a massive downregulation of enzymes belonging to the glycolytic pathway and fatty acids degradation, strongly suggesting a decrease of energy production. A strong reduction in the expression of the N-acetylneuraminate lyases (NanA involved in the sialic acid metabolism was also observed. Using a null NanA mutant and DANA, a substrate analogue acting as competitive inhibitor, we demonstrated that down regulation of NanA affects biofilm formation and adhesion properties of E. coli MV1161. Exposure to alkylating agents also decreased biofilm formation and bacterial adhesion to Caco-2 eukaryotic cell line by the adherent invasive E. coli (AIEC strain LF82. Our data showed that methylation stress impairs E. coli adhesion properties and suggest a possible role of NanA in biofilm formation and bacteria host interactions.

  20. Exposure of E. coli to DNA-Methylating Agents Impairs Biofilm Formation and Invasion of Eukaryotic Cells via Down Regulation of the N-Acetylneuraminate Lyase NanA.

    Science.gov (United States)

    Di Pasquale, Pamela; Caterino, Marianna; Di Somma, Angela; Squillace, Marta; Rossi, Elio; Landini, Paolo; Iebba, Valerio; Schippa, Serena; Papa, Rosanna; Selan, Laura; Artini, Marco; Palamara, Anna Teresa; Palamara, Annateresa; Duilio, Angela

    2016-01-01

    DNA methylation damage can be induced by endogenous and exogenous chemical agents, which has led every living organism to develop suitable response strategies. We investigated protein expression profiles of Escherichia coli upon exposure to the alkylating agent methyl-methane sulfonate (MMS) by differential proteomics. Quantitative proteomic data showed a massive downregulation of enzymes belonging to the glycolytic pathway and fatty acids degradation, strongly suggesting a decrease of energy production. A strong reduction in the expression of the N-acetylneuraminate lyases (NanA) involved in the sialic acid metabolism was also observed. Using a null NanA mutant and DANA, a substrate analog acting as competitive inhibitor, we demonstrated that down regulation of NanA affects biofilm formation and adhesion properties of E. coli MV1161. Exposure to alkylating agents also decreased biofilm formation and bacterial adhesion to Caco-2 eukaryotic cell line by the adherent invasive E. coli (AIEC) strain LF82. Our data showed that methylation stress impairs E. coli adhesion properties and suggest a possible role of NanA in biofilm formation and bacteria host interactions.

  1. Exposure of E. coli to DNA-Methylating Agents Impairs Biofilm Formation and Invasion of Eukaryotic Cells via Down Regulation of the N-Acetylneuraminate Lyase NanA

    Science.gov (United States)

    Di Pasquale, Pamela; Caterino, Marianna; Di Somma, Angela; Squillace, Marta; Rossi, Elio; Landini, Paolo; Iebba, Valerio; Schippa, Serena; Papa, Rosanna; Selan, Laura; Artini, Marco; Palamara, Anna Teresa; Duilio, Angela

    2016-01-01

    DNA methylation damage can be induced by endogenous and exogenous chemical agents, which has led every living organism to develop suitable response strategies. We investigated protein expression profiles of Escherichia coli upon exposure to the alkylating agent methyl-methane sulfonate (MMS) by differential proteomics. Quantitative proteomic data showed a massive downregulation of enzymes belonging to the glycolytic pathway and fatty acids degradation, strongly suggesting a decrease of energy production. A strong reduction in the expression of the N-acetylneuraminate lyases (NanA) involved in the sialic acid metabolism was also observed. Using a null NanA mutant and DANA, a substrate analog acting as competitive inhibitor, we demonstrated that down regulation of NanA affects biofilm formation and adhesion properties of E. coli MV1161. Exposure to alkylating agents also decreased biofilm formation and bacterial adhesion to Caco-2 eukaryotic cell line by the adherent invasive E. coli (AIEC) strain LF82. Our data showed that methylation stress impairs E. coli adhesion properties and suggest a possible role of NanA in biofilm formation and bacteria host interactions. PMID:26904018

  2. Regulation of osteoprotegerin expression by Notch signaling in human oral squamous cell carcinoma cell line

    Institute of Scientific and Technical Information of China (English)

    Jeeranan Manokawinchoke; Thanaphum Osathanon; Prasit Pavasant

    2016-01-01

    Objective: To investigate the influence of Notch signaling on osteoprotegerin (OPG) expression in a human oral squamous cell carcinoma cell line. Methods: Activation of Notch signaling was performed by seeding cells on Jagged1 immobilized surfaces. In other experiments, a γ-secretase inhibitor was added to the culture medium to inhibit intracellular Notch signaling. OPG mRNA and protein were determined by real-time PCR and ELISA, respectively. Finally, publicly available microarray database analysis was performed using connection up- or down-regulation expression analysis of microarrays software. Results: Jagged1-treatment of HSC-4 cells enhanced HES1 and HEY1 mRNA expres-sion, confirming the intracellular activation of Notch signaling. OPG mRNA and protein levels were significantly suppressed upon Jagged1 treatment. Correspondingly, HSC-4 cells treated with a γ-secretase inhibitor resulted in a significant reduction of HES1 and HEY1 mRNA levels, and a marked increase in OPG protein expression was observed. These results implied that Notch signaling regulated OPG expression in HSC-4 cells. However, Jagged1 did not alter OPG expression in another human oral squamous cell carcinoma cell line (HSC-5) or a human head and neck squamous cell carcinoma cell line (HN22). Conclusions: Notch signaling regulated OPG expression in an HSC-4 cell line and this mechanism could be cell line specific.

  3. Regulation of osteoprotegerin expression by Notch signaling in human oral squamous cell carcinoma cell line

    Institute of Scientific and Technical Information of China (English)

    Jeeranan Manokawinchoke; Thanaphum Osathanon; Prasit Pavasant

    2016-01-01

    Objective: To investigate the influence of Notch signaling on osteoprotegerin(OPG)expression in a human oral squamous cell carcinoma cell line.Methods: Activation of Notch signaling was performed by seeding cells on Jagged1 immobilized surfaces. In other experiments, a g-secretase inhibitor was added to the culture medium to inhibit intracellular Notch signaling. OPG m RNA and protein were determined by real-time PCR and ELISA, respectively. Finally, publicly available microarray database analysis was performed using connection up- or down-regulation expression analysis of microarrays software.Results: Jagged1-treatment of HSC-4 cells enhanced HES1 and HEY1 m RNA expression, confirming the intracellular activation of Notch signaling. OPG m RNA and protein levels were significantly suppressed upon Jagged1 treatment. Correspondingly, HSC-4 cells treated with a g-secretase inhibitor resulted in a significant reduction of HES1 and HEY1 m RNA levels, and a marked increase in OPG protein expression was observed.These results implied that Notch signaling regulated OPG expression in HSC-4 cells.However, Jagged1 did not alter OPG expression in another human oral squamous cell carcinoma cell line(HSC-5) or a human head and neck squamous cell carcinoma cell line(HN22).Conclusions: Notch signaling regulated OPG expression in an HSC-4 cell line and this mechanism could be cell line specific.

  4. Triptolide down-regulates COX-2 expression and PGE2 release by suppressing the activity of NF-κB and MAP kinases in lipopolysaccharide-treated PC12 cells.

    Science.gov (United States)

    Geng, Yu; Fang, Marong; Wang, Jing; Yu, Haiyan; Hu, Zhiying; Yew, David T; Chen, Wei

    2012-03-01

    As an active compound extracted from the Chinese herb Tripterygium wilfordii, triptolide (TP) was demonstrated to have potent antiinflammatory and immunosuppressive properties in previous studies. Recently, it has been shown that TP prevented the loss of dopaminergic neurons in the substantia nigra of rats in a model of Parkinson's disease, but little is known about the precise neuroprotective mechanism of TP. This study was designed to elucidate whether the neuroprotective effect of TP is partially based on its direct inhibition of inflammatory molecules by investigating the effects of TP on the expression of cyclooxygenase (COX)-2 and prostaglandin E2 (PGE2) related to the nuclear factor (NF)-κB pathway in lipopolysaccharide (LPS)-stimulated PC12 cells. The activation of related upstream molecules such as NF-κB, P38, extracellular signal-regulated kinase (ERK)1/2, and beta-alanyl-alpha-ketoglutarate transaminase (AKT), in PC12 cells were investigated by real time polymerase chain reaction (PCR), western blotting and enzyme-linked immunosorbent assay (ELISA). Our results showed that TP directly inhibited the expression of both mRNA and protein of COX-2 (p < 0.01), decreased PGE2 production (p < 0.01) in a dose-dependent manner, down-regulated NF-κB activity (p < 0.01), and significantly inhibited the phosphorylation of p38, ERK1/2 (p42/p44) and AKT in PC12 cells after LPS challenge. This suggests that the neuroprotective effects of TP may be partially mediated by direct inhibition of the expression of COX-2, activation of NF-κB, and phosphorylation of p38, ERK1/2 (p42/p44) and AKT proteins of neuronal cells.

  5. Down-regulation of histone deacetylase 4, -5 and -6 as a mechanism of synergistic enhancement of apoptosis in human lung cancer cells treated with the combination of a synthetic retinoid, Am80 and green tea catechin.

    Science.gov (United States)

    Oya, Yukiko; Mondal, Anupom; Rawangkan, Anchalee; Umsumarng, Sonthaya; Iida, Keisuke; Watanabe, Tatsuro; Kanno, Miki; Suzuki, Kaori; Li, Zhenghao; Kagechika, Hiroyuki; Shudo, Koichi; Fujiki, Hirota; Suganuma, Masami

    2017-04-01

    (-)-Epigallocatechin gallate (EGCG), a green tea catechin, acts as a synergist with various anticancer drugs, including retinoids. Am80 is a synthetic retinoid with a different structure from all-trans-retinoic acid: Am80 is now clinically utilized as a new drug for relapsed and intractable acute promyelocytic leukemia patients. Our experiments showed that the combination of EGCG and Am80 synergistically induced both apoptosis in human lung cancer cell line PC-9 and up-regulated expressions of growth arrest and DNA damage-inducible gene 153 (GADD153), death receptor 5, and p21(waf1) genes in the cells. To understand the mechanisms of synergistic anticancer activity of the combination, we gave special attention to the lysine acetylation of proteins. Proteomic analysis using nanoLC-ESI-MS/MS revealed that PC-9 cells treated with the combination contained 331 acetylated proteins, while nontreated cells contained 553 acetylated proteins, and 59 acetylated proteins were found in both groups. Among them, the combination increased acetylated-p53 and acetylated-α-tubulin through reduction of histone deacetylase (HDAC) activity in cytosol fraction, although the levels of acetylation in histones H3 or H4 did not change, and the combination reduced protein levels of HDAC4, -5 and -6 by 20% to 80%. Moreover, we found that a specific inhibitor of HDAC4 and -5 strongly induced p21(waf1) gene expression, and that of HDAC6 induced both GADD153 and p21(waf1) gene expression, which resulted in apoptosis. All results demonstrate that EGCG in combination with Am80 changes levels of acetylation in nonhistone proteins via down-regulation of HDAC4, -5 and -6 and stimulates apoptotic induction.

  6. Down-regulation of CEACAM1 in breast cancer.

    Science.gov (United States)

    Yang, Changcheng; He, Pingqing; Liu, Yiwen; He, Yiqing; Yang, Cuixia; Du, Yan; Zhou, Muqing; Wang, Wenjuan; Zhang, Guoliang; Wu, Man; Gao, Feng

    2015-10-01

    Carcinoembryonic antigen-related adhesion molecule 1 (CEACAM1) is a type 1 transmembrane glycoprotein belonging to the CEA family, which has been found to exist as either soluble forms in body fluids or membrane-bound forms on the cell surface. Aberrant CEACAM1 expression is associated with tumor progression and has been found in a variety of human malignancies. Increasing interest has been devoted to the expression of CEACAM1 in breast cancer, but most of these findings are contradictory. The aim of this study was to investigate CEACAM1 expression in breast cancer in greater detail. Using immunohistochemical staining, we found that CEACAM1 expression was reduced or lost in breast cancer tissues compared with noncancerous breast tissues. In addition, soluble CEACAM1 levels in the culture medium of breast cancer cell lines were significantly lower than those in a nontumorigenic breast epithelial cell line. Immunofluorescence analysis consistently showed that breast cancer cell lines have relatively low expression of membrane-bound CEACAM1. Furthermore, CEACAM1 mRNA and protein expression levels were down-regulated in breast cancer cell lines as measured using real-time reverse transcriptase-polymerase chain reaction and western blot analysis, respectively. Taken together, our results demonstrate a systematic down-regulation of CEACAM1 in breast cancer and suggest that a strategy to restore CEACAM1 expression may be helpful for the treatment of breast cancer.

  7. Caffeine induces matrix metalloproteinase-2 (MMP-2) and MMP-9 down-regulation in human leukemia U937 cells via Ca2+/ROS-mediated suppression of ERK/c-fos pathway and activation of p38 MAPK/c-jun pathway.

    Science.gov (United States)

    Liu, Wen-Hsin; Chang, Long-Sen

    2010-09-01

    Caffeine attenuated invasion of human leukemia U937 cells with characteristic of decreased protein expression and mRNA levels of matrix metalloproteinase-2 (MMP-2) and MMP-9. Down-regulation of MMP-2 and MMP-9 in U937 cells was abrogated by abolishment of caffeine-elicited increase in intracellular Ca(2+) concentration and ROS generation. Pretreatment with BAPTA-AM (Ca(2+) chelator) and N-acetylcysteine (ROS scavenger) abolished caffeine-induced ERK inactivation and p38 MPAK activation. Moreover, caffeine treatment led to MAPK phosphatase-1 (MKP-1) down-regulation and protein phosphatase 2A catalytic subunit (PP2Ac) up-regulation, which were involved in cross-talk between p38 MAPK and ERK. Transfection of constitutively active MEK1 or pretreatment with SB202190 (p38 MAPK inhibitor) restored MMP-2 and MMP-9 protein expression in caffeine-treated cells. Caffeine treatment repressed ERK-mediated c-Fos phosphorylation but evoked p38 MAPK-mediated c-Jun phosphorylation. Knock-down of c-Fos and c-Jun by siRNA reflected that c-Fos counteracted the effect of c-Jun on MMP-2/MMP-9 down-regulation. Taken together, our data indicate that MMP-2/MMP-9 down-regulation in caffeine-treated U937 cells is elicited by Ca(2+)/ROS-mediated suppression of ERK/c-Fos pathway and activation of p38 MAPK/c-Jun pathway.

  8. Expression of cell wall related genes in basal and ear internodes of silking brown-midrib-3, caffeic acid O-methyltransferase (COMT down-regulated, and normal maize plants

    Directory of Open Access Journals (Sweden)

    Martinant Jean-Pierre

    2008-06-01

    Full Text Available Abstract Background Silage maize is a major forage and energy resource for cattle feeding, and several studies have shown that lignin content and structure are the determining factors in forage maize feeding value. In maize, four natural brown-midrib mutants have modified lignin content, lignin structure and cell wall digestibility. The greatest lignin reduction and the highest cell wall digestibility were observed in the brown-midrib-3 (bm3 mutant, which is disrupted in the caffeic acid O-methyltransferase (COMT gene. Results Expression of cell wall related genes was investigated in basal and ear internodes of normal, COMT antisens (AS225, and bm3 maize plants of the INRA F2 line. A cell wall macro-array was developed with 651 gene specific tags of genes specifically involved in cell wall biogenesis. When comparing basal (older lignifying and ear (younger lignifying internodes of the normal line, all genes known to be involved in constitutive monolignol biosynthesis had a higher expression in younger ear internodes. The expression of the COMT gene was heavily reduced, especially in the younger lignifying tissues of the ear internode. Despite the fact that AS225 transgene expression was driven only in sclerenchyma tissues, COMT expression was also heavily reduced in AS225 ear and basal internodes. COMT disruption or down-regulation led to differential expressions of a few lignin pathway genes, which were all over-expressed, except for a phenylalanine ammonia-lyase gene. More unexpectedly, several transcription factor genes, cell signaling genes, transport and detoxification genes, genes involved in cell wall carbohydrate metabolism and genes encoding cell wall proteins, were differentially expressed, and mostly over-expressed, in COMT-deficient plants. Conclusion Differential gene expressions in COMT-deficient plants highlighted a probable disturbance in cell wall assembly. In addition, the gene expressions suggested modified chronology of the

  9. Merkel cell carcinoma: case report.

    Science.gov (United States)

    Sustić, Nela; Biljan, Darko; Orkić, Zelimir; Lizatović, Dario; Milas-Ahić, Jasminka

    2010-04-01

    Merkel cell carcinoma (MCC) is a rare, aggressive neuroendocrine carcinoma of the skin. Although it is 40 times less common than malignant melanoma, its mortality is much higher compared to melanoma. From 1986 to 2001 there was rapidly increasing incidence in reported cases of MCC, with a tripling in the rate over this 15-year period. The vast majority of MCC presents on sun-exposed skin. The head and neck area is the most common site of tumor occurrence. We present 70-year old female patient with painless red-colored nodule, size 2 x 2 x 2 cm on the dorsal side of mid left forearm. The surgical excision with negative margins was performed, and pathohistological analysis confirmed Merkel cell carcinoma. Sentinel lymph node biopsy was negative. In conclusion, as MCC is a very aggressive rare skin carcinoma with lethal outcome, it should be mandatory to perform biopsies of any suspected skin lesion.

  10. Depletion of human micro-RNA miR-125b reveals that it is critical for the proliferation of differentiated cells but not for the down-regulation of putative targets during differentiation.

    Science.gov (United States)

    Lee, Yong Sun; Kim, Hak Kyun; Chung, Sangmi; Kim, Kwang-Soo; Dutta, Anindya

    2005-04-29

    Micro-RNAs are small non-coding RNAs that regulate target gene expression post-transcriptionally through base pairing with the target messenger RNA. Functional characterization of micro-RNAs awaits robust experimental methods to knock-down a micro-RNA as well as to assay its function in vivo. In addition to the recently developed method to sequester micro-RNA with 2'-O-methyl antisense oligonucleotide, we report that small interfering RNA against the loop region of a micro-RNA precursor can be used to deplete the micro-RNA. The depletion of miR-125b by this method had a profound effect on the proliferation of adult differentiated cancer cells, and this proliferation defect was rescued by co-transfected mature micro-RNA. This technique has unique advantages over the 2'-O-methyl antisense oligonucleotide and can be used to determine micro-RNA function, assay micro-RNAs in vivo, and identify the contribution of a predicted micro-RNA precursor to the pool of mature micro-RNA in a given cell. miR-125b and let-7 micro-RNAs are induced, whereas their putative targets, lin-28 and lin-41, are decreased during in vitro differentiation of Tera-2 or embryonic stem cells. Experimental increase or decrease of micro-RNA concentrations did not, however, affect the levels of the targets, a finding that is explained by the fact that the down-regulation of the targets appears to be mostly at the transcriptional level in these in vitro differentiation systems. Collectively these results reveal the importance of micro-RNA depletion strategies for directly determining micro-RNA function in vivo.

  11. Simultaneous Laryngeal Squamous Cell Carcinoma and Papillary Thyroid Carcinoma

    Directory of Open Access Journals (Sweden)

    Bighan Khademi

    2011-04-01

    Full Text Available The association of squamous cell carcinoma of the larynx with thyroid papillary carcinoma is an unusual finding. From 2004 to 2011, approximately 250 patients underwent laryngectomies due to squamous cell carcinoma of the larynx at the Otolaryngology Department of Khalili Hospital, affiliated with Shiraz University of Medical Sciences, Shiraz, Iran. In three patients, synchronous occurrence of squamous cell carcinoma and thyroid papillary carcinoma was found. Histopathologic study of the lymph nodes revealed metastatic papillary thyroid carcinoma in one case. We report three cases of thyroid papillary carcinoma incidentally found on histological examinations of resected thyroid lobes, as a procedure required for treatment of head and neck squamous cell carcinoma. In comparison, laryngeal squamous cell carcinoma needs more aggressive treatment than well-differentiated thyroid carcinoma. The prevalence of thyroid papillary carcinoma, as an incidental finding in our study was 0.01%. Therefore, preoperative evaluation of the thyroid gland by ultrasonography and fine needle aspiration biopsy of suspicious lesions is recommended in patients who are candidates for open laryngectomy.

  12. Multidrug resistance reversal in human gastric carcinoma cells by neferine

    Institute of Scientific and Technical Information of China (English)

    Jian-Guo Cao; Xiao-Qing Tang; Shu-Hong Shi

    2004-01-01

    AIM: To investigate the reversal effect of neferine on multidrug resistance in human gastric carcinoma cell line.METHODS: Cells of a human gastric cancer cells line, SGC7901,and its vincristine (VCR) -resistant variant, SGC7901/VCR,were cultivated with or without neferine and/or VCR. The cytotoxic effect of VCR was evaluated by the MTT assay. Cell apoptosis induced by VCR was determined by flow cytometry (FCM). The expression of P-glycoprotein (P-gp) and a multidrug-resistance-associated protein (MRP) in cells was examined by immunofluorescence and FCM.RESULTS: Neferine at the concentration from 2.5 μmol/L to 10 μmol/L had no cytotoxicity to SGC7901 cells, and its variant SGC7901/VCR cells. The IC50 of VCR against SGC7901 and SGC7901/VCR cells was 0.059 μg/mL and 2.32 μg/mL,respectively, indicating that SGC7901/VCR cells were 39 times more resistant to VCR than its parent SGC7 901 cells. After treatment with neferine at concentrations of 2.5, 5 and 10 μmol/L, the IC50 of VCR to SGC7901/VCR cell line decreased to 0.340, 0.128 and 0.053 μg/mL, respectively,thus, increased the chemosensitivity by 6.8-, 18.1- and 43.8-fold, respectively. SGC7901/VCR cells were apoptosis resistant to VCR. Neferine (2.5, 5 and 10 μmol/L) promoted the VCR-induced apoptosis of SGC7901/VCR cells in a dosedependent manner. The expressions of P-gp and MRP were strongly positive in SGC7901/VCR cells, which were significantly down-regulated after treatment with neferine (10 μmol/L)for 24 h.CONCLUSION: Neferine reverses multidrug resistance of human gastric carcinoma SGC7901/VCR cells, which may be associated with the down-regulations of P-gp and MRP expression in SGC701/VCR cells.

  13. H9N2 Avian Influenza Virus Protein PB1 Enhances the Immune Responses of Bone Marrow-Derived Dendritic Cells by Down-Regulating miR375

    Science.gov (United States)

    Lin, Jian; Xia, Jing; Tu, Chong Z.; Zhang, Ke Y.; Zeng, Yan; Yang, Qian

    2017-01-01

    Polymerase basic protein 1 (PB1), the catalytic core of the influenza A virus RNA polymerase complex, is essential for viral transcription and replication. Dendritic cells (DCs) possess important antigen presenting ability and a crucial role in recognizing and clearing virus. MicroRNA (miRNA) influence the development of DCs and their ability to present antigens as well as the ability of avian influenza virus (AIV) to infect host cells and replicate. Here, we studied the molecular mechanism underlying the miRNA-mediated regulation of immune function in mouse DCs. We first screened for and verified the induction of miRNAs in DCs after PB1 transfection. Results showed that the viral protein PB1 down-regulated the expression of miR375, miR146, miR339, and miR679 in DCs, consistent with the results of H9N2 virus treatment; however, the expression of miR222 and miR499, also reduced in the presence of PB1, was in contrast to the results of H9N2 virus treatment. Our results suggest that PB1 enhanced the ability of DCs to present antigens, activate lymphocytes, and secrete cytokines, while miR375 over-expression repressed activation of DC maturation. Nevertheless, PB1 could not promote DC maturation once miR375 was inhibited. Finally, we revealed that PB1 inhibited the P-Jnk/Jnk signaling pathway, but activated the p-Erk/Erk signaling pathway. While inhibition of miR375 -activated the p-Erk/Erk and p-p38/p38 signaling pathway, but repressed the P-Jnk/Jnk signaling pathway. Taken together, results of our studies shed new light on the roles and mechanisms of PB1 and miR375 in regulating DC function and suggest new strategies for combating AIV.

  14. Stages of Merkel Cell Carcinoma

    Science.gov (United States)

    ... when Merkel cells grow out of control. Merkel cell carcinoma starts most often in areas of skin exposed to the sun, especially the head and neck, as well as the arms, legs, and trunk. Enlarge Anatomy of the skin showing the epidermis, ...

  15. Sorafenib in renal cell carcinoma.

    Science.gov (United States)

    Davoudi, Ehsan Taghizadeh; bin-Noordin, Mohamed Ibrahim; Javar, Hamid Akbari; Kadivar, Ali; Sabeti, Bahare

    2014-01-01

    Cancer is among most important causes of death in recent decades. Whoever the renal cell carcinoma incidence is low but it seems it is more complicated than the other cancers in terms of pathophysiology and treatments. The purpose of this work is to provide an overview and also deeper insight to renal cell carcinoma and the steps which have been taken to reach more specific treatment and target therapy, in this type of cancer by developing most effective agents such as Sorafenib. To achieve this goal hundreds of research paper and published work has been overviewed and due to limitation of space in a paper just focus in most important points on renal cell carcinoma, treatment of RCC and clinical development of Sorafenib. The information presented this paper shows the advanced of human knowledge to provide more efficient drug in treatment of some complicated cancer such as RCC in promising much better future to fight killing disease.

  16. 左炔诺孕酮下调survivin基因促进人子宫肌瘤细胞凋亡%Levonorgestrel induces apoptosis through down-regulation of survivin gene expression in human uterine leiomyoma cells

    Institute of Scientific and Technical Information of China (English)

    仇黎丽; 徐青; 徐昌芬

    2012-01-01

    Objective To explore the function of Survivin in levonorgestrel ( LNG) -induced apoptosis in cultured human uterine leiomyoma cells (UtLMC) in vitro. Methods AO/EB double staining was applied to discriminate the apoptotic cells from dead ones. The changes of survivin mRNA expressions in UtLMC after LNG treatment were detected by semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) method. Furthermore, Western blot was used to determine survivin protein expression in LNG-induced UtLMC apoptosis. Results The result of AO/EB double staining showed that there were more viable apoptotic cells in LNG-treated groups than control group and the number of non-viable apoptotic cells and dead cells increased while the concentration of LNG elevated. Survivin expression in LNG-treated cells was decreased both at mRNA and protein levels according to RT-PCR and Western blot results. Conclusions Levonorgestrel treatment may induce a substantial apoptotic response in UtLMC, which was enhanced with the raise of levonorgestrel concentration. It may be due to the marked down-regulation of survivin gene expression.%目的 探讨左炔诺孕酮(LNG)诱导人子宫肌瘤细胞(UtLMC)凋亡过程中survivin的表达变化.方法 原代培养人子宫肌瘤细胞传代后,加入不同浓度LNG,以AO/EB双染法区分早、晚期凋亡细胞和坏死细胞;RT-PCR检测抑凋亡基因survivin mRNA的表达,Western blot测定survivin蛋白的表达.结果 10 mg/L LNG作用UtLMC后早期凋亡细胞多于阴性对照组,随着LNG剂量的增加,晚期凋亡细胞也逐渐增多,核浓聚、偏位,被染成桔红色;在10 mg/L以上LNG诱导的UtLMC凋亡中,survivin mRNA表达显著下降,蛋白表达由对照组的33.82±0.02下降至10.37±0.03 (P <0.05).结论 一定浓度的左炔诺孕酮所诱导的人子宫肌瘤细胞凋亡,可能与抑制survivin抗凋亡活性相关.

  17. EMP INDUCES APOPTOSIS IN HUMAN LUNG CARCINOMA CELL LINE GLC-82

    Institute of Scientific and Technical Information of China (English)

    曹晓哲; 赵梅兰; 王德文; 董波

    2002-01-01

    Objective: To study the effect of electromagnetic pulse (EMP) on apoptosis of human lung carcinoma cell line GLC-82. Methods: The injury changes in GLC-82 cells after irradiated by EMP (electric field intensity was 60 kV/m with 5 pulses/2 min) were analyzed by cytometry, MTT chronometry and flow cytometry. The immuno- histochemical SP staining was used to determine the expressions of bcl-2 protein and p53 protein. The stained positive cells were analyzed by CMIAS-II image analysis system at a magnification 400. All data were analyzed by SPSS8.0 software. Results: EMP could obviously inhibited lung carcinoma cell line GLC-82 proliferation and increased the number of non-adherent cells. The absorbance value (A570) of MTT decreased immediately, at 0 h, 1 h and 6 h after the GLC-82 cells irradiated by EMP as compared with control group. The highest apoptosis rate was found to reach 13.38% by flow cytometry at 6 h after EMP irradiation. Down-regulation of bcl-2 expression and up-regulation of bax expression were induced by EMP. Conclusion: EMP promoted apoptosis of GLC-82 cells. At same time, EMP can down-regulate bcl-2 expression and up-regulate p53 expression in GLC-82 cells. The bcl-2 and the p53 protein may involve the apoptotic process.

  18. Potential targets for lung squamous cell carcinoma

    Science.gov (United States)

    Researchers have identified potential therapeutic targets in lung squamous cell carcinoma, the second most common form of lung cancer. The Cancer Genome Atlas (TCGA) Research Network study comprehensively characterized the lung squamous cell carcinoma gen

  19. Metastatic basal cell carcinoma caused by carcinoma misdiagnosed as acne - case report and literature review

    DEFF Research Database (Denmark)

    Aydin, Dogu; Hölmich, Lisbet Rosenkrantz; Jakobsen, Linda P

    2016-01-01

    Basal cell carcinoma can be misdiagnosed as acne; thus, carcinoma should be considered in treatment-resistant acne. Although rare, neglected basal cell carcinoma increases the risk of metastasis.......Basal cell carcinoma can be misdiagnosed as acne; thus, carcinoma should be considered in treatment-resistant acne. Although rare, neglected basal cell carcinoma increases the risk of metastasis....

  20. Primary orbital squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Ana L. Campos Arbulú

    2017-02-01

    Full Text Available Primary orbital squamous cell carcinoma is a rare entity. There is little published literature. We report a case of primary squamous cell carcinoma of the orbital soft tissues. Surgical resection offered the best treatment for the patient. Complete resection of the lesion was achieved. The patient received adjuvant radiotherapy due to the proximity of the lesion to the surgical margins. Surgical treatment is feasible and should be considered as part of the surgeon's arsenal. However, therapeutic decisions must be made on a case-by-case basis

  1. [The Dutch guideline 'Renal cell carcinoma'].

    NARCIS (Netherlands)

    Osanto, S.; Bex, A.; Hulsbergen- van de Kaa, C.A.; Soetekouw, P.M.M.B.; Stemkens, D.

    2012-01-01

    The Dutch guideline 'Renal Cell Carcinoma' has been revised on the basis of new literature. With the assistance of the Netherlands Cancer Registry an assessment was made of the current care for patients with renal cell carcinoma. Renal cell carcinoma is a type of cancer for which knowledge of the ge

  2. Stem cell pluripotency factor NANOG is expressed in human fetal gonocytes, testicular carcinoma in situ and germ cell tumours

    DEFF Research Database (Denmark)

    Hoei-Hansen, C E; Almstrup, K; Nielsen, J E

    2005-01-01

    AIMS: NANOG is a key regulator of embryonic stem cell (ESC) self-renewal and pluripotency. Our recent genome-wide gene expression profiling study of the precursor of testicular germ cell tumours, carcinoma in situ testis (CIS), showed close similarity between ESC and CIS, including high NANOG...... earlier than for OCT-4. We detected no expression at the protein level in normal testis. CONCLUSIONS: NANOG is a new marker for testicular CIS and germ cell tumours and the high level of NANOG along with OCT-4 are determinants of the stem cell-like pluripotency of the preinvasive CIS cell. Timing of NANOG......; seminoma and embryonal carcinoma were strongly positive, differentiated somatic elements of teratoma were negative. We provide evidence for the fetal origin of testicular cancer as we detected strong expression of NANOG in fetal gonocytes up to gestational week 20, with subsequent down-regulation occurring...

  3. Spontaneous regression of metastatic Merkel cell carcinoma.

    LENUS (Irish Health Repository)

    Hassan, S J

    2010-01-01

    Merkel cell carcinoma is a rare aggressive neuroendocrine carcinoma of the skin predominantly affecting elderly Caucasians. It has a high rate of local recurrence and regional lymph node metastases. It is associated with a poor prognosis. Complete spontaneous regression of Merkel cell carcinoma has been reported but is a poorly understood phenomenon. Here we present a case of complete spontaneous regression of metastatic Merkel cell carcinoma demonstrating a markedly different pattern of events from those previously published.

  4. Merkel cell carcinoma: a review.

    Science.gov (United States)

    Oram, Christian W; Bartus, Cynthia L; Purcell, Stephen M

    2016-04-01

    Merkel cell carcinoma (MCC) is a rare neuroendocrine tumor of unknown origin that usually presents in the elderly population. A novel polyomavirus has been associated with a large percentage of tumors. Immune response plays an important role in pathogenesis of MCC. This article reviews the history, pathogenesis, presentation, and treatment of MCC. Future treatments also are discussed briefly.

  5. Cryotherapy in basal cell carcinoma

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    Sandra A

    1999-01-01

    Full Text Available Cryotherapy has proved to be an effective tool in the management of various dermatoses. We report 6 patients with histopathologically proven basal cell carcinoma of variable sizes treated with liquid nitrogen cryotherapy by the open spray technique. Lesions tended to heal with depigmentation and scar formation. However depigmented areas often repigmented over a period of time.

  6. Study on immune function of dendritic cells in patients with esophageal carcinoma

    Institute of Scientific and Technical Information of China (English)

    Shen-Ren Chen; Yi-Ping Luo; Jin-Kun Zhang; Wei Yang; Zhi-Chao Zhen; Lin-Xin Chen; Wei Zhang

    2004-01-01

    AIM: To investigate the immune function of dendritic cells from both peripheral blood and operated tissues of esophageal carcinoma patients in order to find the relationship between the immune function of dendritic cells and the pathogenesis of esophageal carcinoma.METHODS: The expression of CD83, CD80, and CD86 on the surface of dendritic cells cultured from the peripheral blood of patients was detected compared with that from health donors using flow cytometry. The ability of dendritic cells to induce T lymphocyte proliferation was evaluated by a liquid scintillation counter. The expression of CD80, CD86,CD83, and S-100 proteins was assessed in esophageal carcinoma tissues using immunohistochemical method.RESULTS: Compared with those from healthy donors,dendirtic cells cultured from the peripheral blood of patients expressed lower CD80 and CD86. Furthermore, the ability of dendritic cells in patients to induce T lymphocyte proliferation was significantly lower than that of the control group. Compared with the control group, the positive expression ratio and frequencies of CDS0, CD86, and S100 in esophageal carcinoma tissues were significantly down regulated. The expression of CD83 was up-regulated in the pericancerous tissues, but no expression was found in the cancerous nodules,CONCLUSION: The impaired immune function and the decreased number of dendritic cells cause pathogenesis and progression of esophageal carcinoma.

  7. Metastatic renal cell carcinoma management

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    Flavio L. Heldwein

    2009-06-01

    Full Text Available PURPOSE: To assess the current treatment of metastatic renal cell carcinoma, focusing on medical treatment options. MATERIAL AND METHODS: The most important recent publications have been selected after a literature search employing PubMed using the search terms: advanced and metastatic renal cell carcinoma, anti-angiogenesis drugs and systemic therapy; also significant meeting abstracts were consulted. RESULTS: Progress in understanding the molecular basis of renal cell carcinoma, especially related to genetics and angiogenesis, has been achieved mainly through of the study of von Hippel-Lindau disease. A great variety of active agents have been developed and tested in metastatic renal cell carcinoma (mRCC patients. New specific molecular therapies in metastatic disease are discussed. Sunitinib, Sorafenib and Bevacizumab increase the progression-free survival when compared to therapy with cytokines. Temsirolimus increases overall survival in high-risk patients. Growth factors and regulatory enzymes, such as carbonic anhydrase IX may be targets for future therapies. CONCLUSIONS: A broader knowledge of clear cell carcinoma molecular biology has permitted the beginning of a new era in mRCC therapy. Benefits of these novel agents in terms of progression-free and overall survival have been observed in patients with mRCC, and, in many cases, have become the standard of care. Sunitinib is now considered the new reference first-line treatment for mRCC. Despite all the progress in recent years, complete responses are still very rare. Currently, many important issues regarding the use of these agents in the management of metastatic renal cancer still need to be properly addressed.

  8. Small cell undifferentiated carcinoma in the epididymis

    Institute of Scientific and Technical Information of China (English)

    CHEN Jia-wei; YUAN Lin; Hu Hong-hui

    2005-01-01

    @@ Small cell undifferentiated carcinoma is a special type of tumor which is usually found in the lungs. However, it is very rare in extra pulmonary tissues, especially in epididymis. One case of small cell undifferentiated carcinoma in the right epididymis, with partial differentiation to adenocarcinoma and neuroendocrine carcinoma is reported as follows.

  9. Expression Profile of Metastasis-associated Genes in Esophageal Squamous Cell Carcinoma

    Institute of Scientific and Technical Information of China (English)

    LI Pei; LING Zhiqiang; YANG Hongyan; HUANG Youtian; ZHAO Mingyao; ZHENG Zhimin; DONG Ziming

    2006-01-01

    The differentially expressed genes between esophageal squamous cell carcinoma (ESCC)with or without lymphatic metastasis were investigated by gene chip, and the lymphatic metastasisassociated genes were screened out. Expression array was used to detect the mRNA from both the primary carcinoma and the corresponding esophageal epithelium in 15 cases of human ESCC. The lymphatic metastasis-associated genes were screened by bioinformatics between ESCC with or without lymphatic metastasis. The results showed that 43 (4.85%) genes significantly differed between the ESCC with and without lymphatic metastasis (P<0.05), of which 18(2.03%)were upregulated and 25 (2.82 %) down-regulated. The up-regulated genes were involved in cell adhesion molecules and cell membrane receptors and the down-regulated genes were mostly cell cycle regulators and intracellular signaling molecules. It was suggested that lymphatic metastasis-associated genes were screened by gene chip, which was helpful to understand the molecular mechanism of ESCC lymphatic metastasis and lymphatic metastasis-associated genes might be used as diagnostic markers and therapeutic targets for lymphatic metastasis.

  10. Ku70 acetylation and modulation of c-Myc/ATF4/CHOP signaling axis by SIRT1 inhibition lead to sensitization of HepG2 cells to TRAIL through induction of DR5 and down-regulation of c-FLIP

    DEFF Research Database (Denmark)

    Kim, Mi-Ju; Hong, Kyung-Soo; Kim, Hak-Bong

    2013-01-01

    In this study, we investigated the role of c-Myc/ATF4/CHOP signaling pathway in sensitization of human hepatoma HepG2 cells to TRAIL. Knockdown of SIRT1 or treatment with SIRT1 inhibitor caused the up-regulation of DR5 and down-regulation of c-FLIP through modulation of c-Myc/ATF4/CHOP pathway, a...

  11. Bax Inhibitor-1 down-regulation in the progression of chronic liver diseases

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    Burra Patrizia

    2010-04-01

    Full Text Available Abstract Background Bax inhibitor-1 (BI-1 is an evolutionary conserved endoplasmic reticulum protein that, when overexpressed in mammalian cells, suppresses the apoptosis induced by Bax, a pro-apoptotic member of the Bcl-2 family. The aims of this study were: (1 to clarify the role of intrinsic anti- and pro-apoptotic mediators, evaluating Bax and BI-1 mRNA and protein expressions in liver tissues from patients with different degrees of liver damage; (2 to determine whether HCV and HBV infections modulate said expression. Methods We examined 62 patients: 39 with chronic hepatitis (CH (31 HCV-related and 8 HBV-related; 7 with cirrhosis (6 HCV-related and 1 HBV-related; 13 with hepatocellular carcinoma (HCC [7 in viral cirrhosis (6 HCV- and 1 HBV-related, 6 in non-viral cirrhosis]; and 3 controls. Bax and BI-1 mRNAs were quantified by real-time PCR, and BI-1 protein expression by Western blot. Results CH tissues expressed significantly higher BI-1 mRNA levels than cirrhotic tissues surrounding HCC (P Conclusions BI-1 expression is down-regulated as liver damage progresses. The high BI-1 mRNAs levels observed in early liver disease may protect virus-infected cells against apoptosis, while their progressive downregulation may facilitate hepatocellular carcinogenesis. HCV genotype seems to have a relevant role in Bax transcript expression.

  12. Gastric Large Cell Neuroendocrine Carcinoma

    Science.gov (United States)

    Rustagi, Tarun; Alekshun, Todd J.

    2010-01-01

    Case: A 63-year-old male presented with unintentional weight loss of 20 pounds over a 4-month duration. He reported loss of appetite, intermittent post-prandial nausea, bloating and early satiety. He also complained of dyspepsia and had been treated for reflux during the previous 2 years. He denied vomiting, dysphagia, odynophagia, abdominal pain, melena, hematochezia, or alterations in bowel habits. Additionally, he denied fevers, night sweats, cough, or dyspnea. He quit smoking 25 years ago, and denied alcohol use. His past medical history was significant for basal cell carcinoma treated with local curative therapy and he was without recurrence on surveillance. Pertinent family history included a paternal uncle with lung cancer at the age of 74. Physical examination was unremarkable except for occult heme-positive stools. Laboratory evaluation revealed elevated liver enzymes (ALT-112, AST-81, AlkPhos-364). CT scan of the chest, abdomen and pelvis showed diffuse heterogeneous liver with extensive nodularity, raising the concern for metastases. Serum tumor-markers: PSA, CEA, CA 19-9, and AFP were all within normal limits. Screening colonoscopy was normal, but esophagogastroduodenoscopy revealed a malignant-appearing ulcerative lesion involving the gastro-esophageal junction and gastric cardia. Pathology confirmed an invasive gastric large cell neuroendocrine carcinoma. Ultrasound-guided fine needle aspiration of a hepatic lesion revealed malignant cells with cytologic features consistent with large-cell type carcinoma and positive immunostaining for synaptophysin favoring neuroendocrine differentiation. A PET-CT demonstrated intense diffuse FDG uptake of the liver, suggesting diffuse hepatic parenchymal infiltration by tumor. There were multiple foci of intense osseous FDG uptake with corresponding osteolytic lesions seen on CT scan. The remaining intra-abdominal and intra-thoracic structures were unremarkable. The patient will receive palliative systemic therapy

  13. FBI-1 enhances ETS-1 signaling activity and promotes proliferation of human colorectal carcinoma cells.

    Science.gov (United States)

    Zhu, Min; Li, Mingyang; Zhang, Fan; Feng, Fan; Chen, Weihao; Yang, Yutao; Cui, Jiajun; Zhang, Dong; Linghu, Enqiang

    2014-01-01

    In this study, we investigated a potential regulatory role of FBI-1 in transcription factor activity of ETS-1. The protein interaction was identified between ETS-1 and FBI-1 in lovo cells. The accumulating data showed that FBI-1 promoted the recruitment of ETS-1 to endogenous promoter of its target genes and increase ETS-1 accumulation in the nuclear. Our work also indicated that the FBI-1 enhances ETS-1 transcription factor activity via down-regulating p53-mediated inhibition on ETS-1. Further, FBI-1 plays a role in regulation of colorectal carcinoma cells proliferation. These findings supported that FBI-1 might be a potential molecule target for treating colorectal carcinoma.

  14. FBI-1 enhances ETS-1 signaling activity and promotes proliferation of human colorectal carcinoma cells.

    Directory of Open Access Journals (Sweden)

    Min Zhu

    Full Text Available In this study, we investigated a potential regulatory role of FBI-1 in transcription factor activity of ETS-1. The protein interaction was identified between ETS-1 and FBI-1 in lovo cells. The accumulating data showed that FBI-1 promoted the recruitment of ETS-1 to endogenous promoter of its target genes and increase ETS-1 accumulation in the nuclear. Our work also indicated that the FBI-1 enhances ETS-1 transcription factor activity via down-regulating p53-mediated inhibition on ETS-1. Further, FBI-1 plays a role in regulation of colorectal carcinoma cells proliferation. These findings supported that FBI-1 might be a potential molecule target for treating colorectal carcinoma.

  15. E3B1/ABI-1 Isoforms Are Down-Regulated in Cancers of Human Gastrointestinal Tract

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    Rafia A. Baba

    2012-01-01

    Full Text Available The expression of E3B1/ABI-1 protein and its role in cancer progression and prognosis are largely unknown in the majority of solid tumors. In this study, we examined the expression pattern of E3B1/ABI-1 protein in histologically confirmed cases of esophageal (squamous cell carcinoma and adenocarcinoma, gastro-esophageal junction, colorectal cancers and corresponding normal tissues freshly resected from a cohort of 135 patients, by Western Blotting and Immunofluorescence Staining. The protein is present in its phosphorylated form in cells and tissues. Depending on the extent of phosphorylation it is either present in hyper-phosphorylated (M. Wt. 72 kDa form or in hypo-phosphorylated form (M. Wt. 68 kDa and 65 kDa. A thorough analysis revealed that expression of E3B1/ABI-1 protein is significantly decreased in esophageal, gastro-esophageal junction and colorectal carcinomas irrespective of age, gender, dietary and smoking habits of the patients. The decrease in expression of E3B1/ABI-1 was consistently observed for all the three isoforms. However, the decrease in the expression of isoforms varied with different forms of cancers. Down-regulation of E3B1/ABI-1 expression in human carcinomas may play a critical role in tumor progression and in determining disease prognosis.

  16. Computer simulation of factors involved in the down-regulation of hormonal effects.

    Science.gov (United States)

    Kurbel, S; Kurbel, B; Dicić, M; Ugraji, V

    1996-03-01

    Down-regulation of hormonal effects is in the presented simulation related to the number of functional receptors and quantity of available hormonal stimulation. The former is in the model substituted with the quantity of stimulation able to produce a full down-regulation (Hs100) of target cells. The halftime (t1/2) of the hormonal effect recovery means the interval before the second hormonal stimulation can elicit half of the initial hormonal effect. Recovered hormonal effects are calculated after periods of two, three, four and five t1/2. The interval among hormonal stimulations varied from 1/2 to 5/2 of t1/2. Shorter than t1/2 intervals showed profound down-regulation even at weak hormonal stimulations (> 20% of Hs100). Stable levels of hormonal effects after frequent hormonal stimulations are found only in cases of very weak stimulations (Hs100). Intervals equalling t1/2 among weak stimulations (Hs100) produced stable hormonal effects. Further prolongation among repeated stimulations improved stability of hormonal effects and even strong stimulations (> 60% of Hs100) were followed with only temporary profound down-regulation. Hormone-binding receptors unable to activate target cells are in the model described as defective. Probability for the target cell to be stimulated is in the model defined as P. Relative quantity of hormonal stimulation per target cell needed to achieve certain P is calculated for cells bearing different proportions of defective receptors. Activation following weak hormone stimulations is highly probable (> 90%) for cells bearing less than 30% of defective receptors. With the proportion of defective receptors over 60%, the activation probability after weak hormone stimulations is reduced (< 66%). Down-regulation can be considered as a modulator of hormonal effects. In prediabetic patients, intense stimulation of pancreatic insulin secretion by frequent or increased ingestion of carbohydrates might lead to sustained hyperinsulinemia. A

  17. Small cell glioblastoma or small cell carcinoma

    DEFF Research Database (Denmark)

    Hilbrandt, Christine; Sathyadas, Sathya; Dahlrot, Rikke H

    2013-01-01

    was admitted to the hospital with left-sided loss of motor function. A MRI revealed a 6 cm tumor in the right temporoparietal area. The histology was consistent with both glioblastoma multiforme (GBM) and small cell lung carcinoma (SCLC) but IHC was suggestive of a SCLC metastasis. PET-CT revealed...

  18. Nevoid basal cell carcinoma syndrome

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    Kannan Karthiga

    2006-01-01

    Full Text Available Binkley and Johnson first reported this syndrome in 1951. But it was in 1960, Gorlin-Goltz established the association of basal cell epithelioma, jaw cyst and bifid ribs, a combination which is now frequently known as Gorlin-Goltz syndrome as well as Nevoid Basal Cell Carcinoma Syndrome (NBCCS. NBCCS is inherited as an autosomal dominant trait with high penetrance and variable expressivity. NBCCS is characterized by variety of cutaneous, dental, osseous, opthalmic, neurologic and sexual abnormalities. One such case of Gorlin-Goltz syndrome is reported here with good illustrations.

  19. Anaplastic giant cell thyroid carcinoma.

    Science.gov (United States)

    Wallin, G; Lundell, G; Tennvall, J

    2004-01-01

    Anaplastic (giant cell) thyroid carcinoma (ATC), is one of the most aggressive malignancies in humans with a median survival time after diagnosis of 3-6 months. Death from ATC was earlier seen because of local growth and suffocation. ATC is uncommon, accounting for less than 5 % of all thyroid carcinomas. The diagnosis can be established by means of multiple fine needle aspiration biopsies, which are neither harmful nor troublesome for the patient. The cytological diagnosis of this high-grade malignant tumour is usually not difficult for a well trained cytologist. The intention to treat patients with ATC is cure, although only few of them survive. The majority of the patients are older than 60 years and treatment must be influenced by their high age. We have by using a combined modality regimen succeeded in achieving local control in most patients. Every effort should be made to control the primary tumour and thereby improve the quality of remaining life and it is important for patients, relatives and the personnel to know that cure is not impossible. Different treatment combinations have been used since 30 years including radiotherapy, cytostatic drugs and surgery, when feasible. In our latest combined regimen, 22 patients were treated with hyper fractionated radiotherapy 1.6Gy x 2 to a total target dose of 46 Gy given preoperatively, 20 mg doxorubicin was administered intravenously once weekly and surgery was carried out 2-3 weeks after the radiotherapy. 17 of these 22 patients were operated upon and none of these 17 patients got a local recurrence. In the future we are awaiting the development of new therapeutic approaches to this aggressive type of carcinoma. Inhibitors of angiogenesis might be useful. Combretastatin has displayed cytotoxicity against ATC cell lines and has had a positive effect on ATC in a patient. Sodium iodide symporter (NIS) genetherapy is also being currently considered for dedifferentiated thyroid carcinomas with the ultimate aim of

  20. EMX2 Is a Predictive Marker for Adjuvant Chemotherapy in Lung Squamous Cell Carcinomas

    Science.gov (United States)

    Zhang, Yi; Tolani, Bhairavi; Mo, Minli; Zhang, Hua; Zheng, Qingfeng; Yang, Yue; Cheng, Runfen; Jin, Joy Q.; Luh, Thomas W.; Yang, Cathryn; Tseng, Hsin-Hui K.; Giroux-Leprieur, Etienne; Woodard, Gavitt A.; Hao, Xishan; Wang, Changli; Jablons, David M.; He, Biao

    2015-01-01

    Background Squamous cell carcinomas (SCC) account for approximately 30% of non-small cell lung cancer (NSCLC). Current staging methods do not adequately predict outcome for this disease. EMX2 is a homeo-domain containing transcription factor known to regulate a key developmental pathway. This study assessed the significance of EMX2 as a prognostic and predictive marker for resectable lung SCC. Methods Two independent cohorts of patients with lung SCC undergoing surgical resection were studied. EMX2 protein expression was examined by immunohistochemistry, Western blot, or immunofluorescence. EMX2 expression levels in tissue specimens were scored and correlated with patient outcomes. Chemo-sensitivity of lung SCC cell lines stably transfected with EMX2 shRNAs to cisplatin, carboplatin, and docetaxel was examined in vitro. Results EMX2 expression was down-regulated in lung SCC tissue samples compared to their matched adjacent normal tissues. Positive EMX2 expression was significantly associated with improved overall survival in stage I lung SCC patients, and in stage II/IIIA lung SCC patients receiving adjuvant chemotherapy. EMX2 expression was also associated with expression of EMT markers in both lung SCC cell lines and tissue samples. Knock-down of EMX2 expression in lung SCC cells promoted chemo-resistance and cell migration. Conclusions EMX2 expression is down-regulated in lung SCC and its down-regulation is associated with chemo-resistance in lung SCC cells, possibly through regulation of Epithelial-to-Mesenchymal Transition (EMT). EMX2 may serve as a novel prognostic marker for stage I lung SCC patients and a prediction marker for stage II/IIIA lung SCC patients receiving adjuvant chemotherapy. PMID:26132438

  1. EMX2 Is a Predictive Marker for Adjuvant Chemotherapy in Lung Squamous Cell Carcinomas.

    Directory of Open Access Journals (Sweden)

    Dongsheng Yue

    Full Text Available Squamous cell carcinomas (SCC account for approximately 30% of non-small cell lung cancer (NSCLC. Current staging methods do not adequately predict outcome for this disease. EMX2 is a homeo-domain containing transcription factor known to regulate a key developmental pathway. This study assessed the significance of EMX2 as a prognostic and predictive marker for resectable lung SCC.Two independent cohorts of patients with lung SCC undergoing surgical resection were studied. EMX2 protein expression was examined by immunohistochemistry, Western blot, or immunofluorescence. EMX2 expression levels in tissue specimens were scored and correlated with patient outcomes. Chemo-sensitivity of lung SCC cell lines stably transfected with EMX2 shRNAs to cisplatin, carboplatin, and docetaxel was examined in vitro.EMX2 expression was down-regulated in lung SCC tissue samples compared to their matched adjacent normal tissues. Positive EMX2 expression was significantly associated with improved overall survival in stage I lung SCC patients, and in stage II/IIIA lung SCC patients receiving adjuvant chemotherapy. EMX2 expression was also associated with expression of EMT markers in both lung SCC cell lines and tissue samples. Knock-down of EMX2 expression in lung SCC cells promoted chemo-resistance and cell migration.EMX2 expression is down-regulated in lung SCC and its down-regulation is associated with chemo-resistance in lung SCC cells, possibly through regulation of Epithelial-to-Mesenchymal Transition (EMT. EMX2 may serve as a novel prognostic marker for stage I lung SCC patients and a prediction marker for stage II/IIIA lung SCC patients receiving adjuvant chemotherapy.

  2. TKTL1 promotes cell proliferation and metastasis in esophageal squamous cell carcinoma.

    Science.gov (United States)

    Li, Juan; Zhu, Shu-Chai; Li, Shu-Guang; Zhao, Yan; Xu, Jin-Rui; Song, Chun-Yang

    2015-08-01

    Transketolase-like-1 (TKTL1), which is a rate-limiting enzyme in the non-oxidative part of the pentose-phosphate pathway, has been demonstrated to promote carcinogenesis through enhanced aerobic glycolysis. Dysregulation of TKTL1 expression also leads to poor prognosis in patients with urothelial and colorectal cancer. However, the expression pattern and underlying cellular functions in human esophageal squamous cell carcinoma (ESCC) remain largely unexplored. In this study, we measured TKTL1 expression in ESCC cell lines and paraffin-embedded ESCC tumor tissues. Our results revealed that TKTL1 expression was upregulated in all of the four ESCC cell lines and in 61.25% (98/160) of ESCC specimens detected, while only 27.5% (11/40) in normal epithelium. Silencing of TKTL1 expression decreased cell proliferation through inhibiting the expression of MKI67 and cyclins including Ccna2, Ccnb1, Ccnd1 and Ccne1. Meanwhile, down-regulation of TKTL1 also associated with increased apoptotic ratio and altered protein expression of Bcl-2 family in ESCC cells. Furthermore, knockdown of TKTL1 significantly reduced the invasive potential of ESCC cells through up-regulation of anti-metastasis genes (MTSS1, TIMP2 and CTSK) and down-regulation of pr-metastasis genes (MMP2, MMP9, MMP10 and MMP13). Taken together, our results indicate that TKTL1 is associated with a more aggressive behavior in ESCC cells and suppresses its expression or enzyme activity might represents a potential target for developing novel therapies in human ESCCs.

  3. Expression of CIDE proteins in clear cell renal cell carcinoma and their prognostic significance.

    Science.gov (United States)

    Yu, Ming; Wang, Hui; Zhao, Jun; Yuan, Yuan; Wang, Chao; Li, Jing; Zhang, Lijun; Zhang, Liying; Li, Qing; Ye, Jing

    2013-06-01

    Clear cell renal cell carcinoma (ccRCC) is the major and aggressive subtype of renal cell carcinoma. It is known to derive its histologic appearance from accumulation of abundant lipids and glycogens. The cell death-inducing DFF45-like effector (CIDE) family has been characterized as the lipid droplet proteins involved in the metabolism of lipid storage droplets. The purpose of this study was to evaluate the expression of CIDE proteins in ccRCC cells and to investigate their prognostic significance. We examined consecutive patients with sporadic ccRCC, who underwent nephrectomy, to measure their mRNA and protein expression of CIDE proteins. We found that Cidec and ADRP expression were significantly up-regulated in ccRCC, compared with normal kidney tissues. Cideb was down-regulated. We also found that Cideb was expressed more in low-grade ccRCC than in high-grade tumors. To further clarify the relationship between Cideb expression and patient prognosis, we evaluated 57 ccRCC patients followed up for 120 months. Reduced ccRCC Cideb expression was associated with a higher Fuhrman nuclear grade. Patients with high Cideb expression had better overall survival rate than those with low expression (p < 0.05). Cideb expression was an independent predictor of survival (p = 0.001). Although the biologic function of Cideb in ccRCC remains unknown, the expression level of Cideb might be a novel predictor of prognosis in ccRCC.

  4. Merkel cell carcinoma versus metastatic small cell primary bronchogenic carcinoma

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    Katya Lisette Velasquez Cantillo

    2013-01-01

    Full Text Available Merkel cell carcinoma (MCC of the skin is a rare, aggressive, malignant neuroendocrine neoplasm. The tumor classically demonstrates positive immunohistochemistry (IHC staining for chromogranin A(ChrA, cytokeratin 20 (CK20, neuron specific enolase (NSE and/or achaete-acute complex-like 1 (MASH1. The newly identified Merkel cell polyomavirus (MCPyV has been found to be associated with most MCC cases. The primary histologic differential diagnoses of cutaneous MCC is small cell primary bronchogenic carcinoma (SCLC; moreover, both are of neuroendocrine origin. SCLC accounts for approximately 10-15% of all primary lung cancer cases; this histologic subtype is a distinct entity with biological and oncological features distinct from non-small cell lung cancer (NSCLC. In contradistinction to MCC, SCLC is classically IHC positive for cytokeratin 7 (CK7 and transcription factor (TTF-1. Similar to SCLC, MCC cell lines may be classified into two different biochemical subgroups designated as Classic and Variant. In our review and case report, we aim to emphasize the importance of a multidisciplinary approach to the approach to this difficult differential diagnosis. We also aim to comment about features of the cells of origin of MCC and SCLC; to summarize the microscopic features of both tumors; and to review their respective epidemiologic, clinical, prognostic and treatment features. We want to emphasize the initial workup study of the differential diagnosis patient, including evaluating clinical lymph nodes, a clinical history of any respiratory abnormality, and chest radiogram. If a diagnosis of primary cutaneous MCC is confirmed, classic treatment includes excision of the primary tumor with wide margins, excision of a sentinel lymph node, and computed tomography, positron emission tomography and/or Fluorine-18-fluorodeoxyglucose positron emission tomography scan studies

  5. Expression of heparanase in basal cell carcinoma and squamous cell carcinoma*

    Science.gov (United States)

    Pinhal, Maria Aparecida Silva; Almeida, Maria Carolina Leal; Costa, Alessandra Scorse; Theodoro, Thérèse Rachell; Serrano, Rodrigo Lorenzetti; Machado Filho, Carlos D'Apparecida Santos

    2016-01-01

    Background Heparanase is an enzyme that cleaves heparan sulfate chains. Oligosaccharides generated by heparanase induce tumor progression. Basal cell carcinoma and squamous cell carcinoma comprise types of nonmelanoma skin cancer. Objectives Evaluate the glycosaminoglycans profile and expression of heparanase in two human cell lines established in culture, immortalized skin keratinocyte (HaCaT) and squamous cell carcinoma (A431) and also investigate the expression of heparanase in basal cell carcinoma, squamous cell carcinoma and eyelid skin of individuals not affected by the disease (control). Methods Glycosaminoglycans were quantified by electrophoresis and indirect ELISA method. The heparanase expression was analyzed by quantitative RT-PCR (qRTPCR). Results The A431 strain showed significant increase in the sulfated glycosaminoglycans, increased heparanase expression and decreased hyaluronic acid, comparing to the HaCaT lineage. The mRNA expression of heparanase was significantly higher in Basal cell carcinoma and squamous cell carcinoma compared with control skin samples. It was also observed increased heparanase expression in squamous cell carcinoma compared to the Basal cell carcinoma. Conclusion The glycosaminoglycans profile, as well as heparanase expression are different between HaCaT and A431 cell lines. The increased expression of heparanase in Basal cell carcinoma and squamous cell carcinoma suggests that this enzyme could be a marker for the diagnosis of such types of non-melanoma cancers, and may be useful as a target molecule for future alternative treatment. PMID:27828631

  6. Apigenin induces autophagic cell death in human papillary thyroid carcinoma BCPAP cells.

    Science.gov (United States)

    Zhang, Li; Cheng, Xian; Gao, Yanyan; Zheng, Jie; Xu, Qiang; Sun, Yang; Guan, Haixia; Yu, Huixin; Sun, Zhen

    2015-11-01

    Apigenin, abundantly present in fruits and vegetables, is recognized as a flavonoid with anti-inflammatory, antioxidant and anticancer properties. In this study, we first investigated the anti-neoplastic effects of apigenin on papillary thyroid carcinoma (PTC) cell line BCPAP cells. Our results show that apigenin inhibited the viability of BCPAP cells in a dose-dependent manner. A large body of evidence demonstrates that autophagy contributes to cell death in certain contexts. In the present study, autophagy was induced by apigenin treatment in BCPAP cells, as evidenced by Beclin-1 accumulation, conversion of LC3 protein, p62 degradation as well as the significantly increased formation of acidic vesicular organelles (AVOs) compared to the control group. 3-MA, an autophagy inhibitor, rescued the cells from apigenin-induced cell death. Notably, apigenin enhanced production of reactive oxygen species (ROS), and subsequent induction of significant DNA damage as monitored by the TUNEL assay. In addition, apigenin treatment caused a significant accumulation of cells in the G2/M phase via down-regulation of Cdc25C expression. Our findings reveal that apigenin inhibits papillary thyroid cancer cell viability by the stimulation of reactive oxygen species (ROS) production, induction of DNA damage, leading to G2/M cell cycle arrest followed by autophagic cell death. Thus, our results provide new insights into the molecular mechanisms underlying apigenin-mediated autophagic cell death and suggest apigenin as a potential chemotherapeutic agent which is able to fight against papillary thyroid cancer.

  7. The CD3 gamma leucine-based receptor-sorting motif is required for efficient ligand-mediated TCR down-regulation

    DEFF Research Database (Denmark)

    von Essen, Marina; Menné, Charlotte; Nielsen, Bodil L;

    2002-01-01

    TCR down-regulation plays an important role in modulating T cell responses both during T cell development and in mature T cells. At least two distinct pathways exist for down-regulation of the TCR. One pathway is activated following TCR ligation and is dependent on tyrosine phosphorylation. The o...

  8. Monocarboxylate transporter 4 facilitates cell proliferation and migration and is associated with poor prognosis in oral squamous cell carcinoma patients.

    Directory of Open Access Journals (Sweden)

    Jiang Zhu

    Full Text Available Monocarboxylate transporter 4 (MCT4 is a cell membrane transporter of lactate. Recent studies have shown that MCT4 is over-expressed in various cancers; however, its role in cancer maintenance and aggressiveness has not been fully demonstrated. This study investigated the role of MCT4 in oral squamous cell carcinoma (OSCC, and found that it is highly expressed in OSCC patients by using immunohistochemistry. Moreover, this over-expression of MCT4 was closely associated with tumor size, TNM classification, lymphatic metastasis, distant metastasis and tumor recurrence, and also poor prognosis. To further study mechanisms of MCT4 in vitro, we used small-interfering RNA to silence its expression in OSCC cell lines. The results showed that knock-down of MCT4 decreased cell proliferation, migration, and invasion. The inhibition of proliferation was associated with down-regulation of p-AKT and p-ERK1/2, while decreased cell migration and invasion may be caused by down-regulation of integrin β4-SRC-FAK and MEK-ERK signaling. Together, these findings provide new insight into the critical role of MCT4 in cell proliferation and metastasis in OSCC.

  9. Spiclomazine Induces Apoptosis Associated with the Suppression of Cell Viability, Migration and Invasion in Pancreatic Carcinoma Cells

    Science.gov (United States)

    Liu, Zuojia; Zheng, Xiliang; Wang, Jin; Wang, Erkang

    2013-01-01

    The effective treatment for pancreatic carcinoma remains critically needed. Herein, this current study showed that spiclomazine treatment caused a reduction in viability in pancreatic carcinoma cell lines CFPAC-1 and MIA PaCa-2 in vitro. It was notable in this regard that, compared with pancreatic carcinoma cells, normal human embryonic kidney (HEK-293) and liver (HL-7702) cells were more resistant to the antigrowth effect of spiclomazine. Biochemically, spiclomazine treatment regulated the expression of protein levels in the apoptosis related pathways. Consistent with this effect, spiclomazine reduced the mitochondria membrane potential, elevated reactive oxygen species, and activated caspase-3/9. In addition, a key finding from this study was that spiclomazine suppressed migration and invasion of cancer cells through down-regulation of MMP-2/9. Collectively, the proposed studies did shed light on the antiproliferation effect of spiclomazine on pancreatic carcinoma cell lines, and further clarified the mechanisms that spiclomazine induced apoptosis associated with the suppression of migration and invasion. PMID:23840452

  10. Spiclomazine induces apoptosis associated with the suppression of cell viability, migration and invasion in pancreatic carcinoma cells.

    Directory of Open Access Journals (Sweden)

    Wenjing Zhao

    Full Text Available The effective treatment for pancreatic carcinoma remains critically needed. Herein, this current study showed that spiclomazine treatment caused a reduction in viability in pancreatic carcinoma cell lines CFPAC-1 and MIA PaCa-2 in vitro. It was notable in this regard that, compared with pancreatic carcinoma cells, normal human embryonic kidney (HEK-293 and liver (HL-7702 cells were more resistant to the antigrowth effect of spiclomazine. Biochemically, spiclomazine treatment regulated the expression of protein levels in the apoptosis related pathways. Consistent with this effect, spiclomazine reduced the mitochondria membrane potential, elevated reactive oxygen species, and activated caspase-3/9. In addition, a key finding from this study was that spiclomazine suppressed migration and invasion of cancer cells through down-regulation of MMP-2/9. Collectively, the proposed studies did shed light on the antiproliferation effect of spiclomazine on pancreatic carcinoma cell lines, and further clarified the mechanisms that spiclomazine induced apoptosis associated with the suppression of migration and invasion.

  11. Basal Cell Carcinoma in The Netherlands

    NARCIS (Netherlands)

    S.C. Flohil (Sophie)

    2012-01-01

    textabstractThere are many different cutaneous malignancies, but malignant melanoma, squamous cell carcinoma (SCC) and basal cell carcinoma (BCC) represent approximately 98% of all skin cancers.In literature, these three skin cancers are often divided into melanoma and nonmelanoma skin cancers (NMSC

  12. Acinar Cell Carcinoma of the Pancreas

    Institute of Scientific and Technical Information of China (English)

    Hua Li; Qiang Li

    2008-01-01

    Acinar cell carcinoma of the pancreas is a rare tumor which is defined as a carcinoma that exhibits pancreatic enzyme production by neoplastic cells. This review includes re-cent developments in our understanding of the epidemiology and pathogenesis of ACC, imaging and pathological diagnosis and ap-proaches to treatment with reference to the literature.

  13. Eyelid Squamous Cell Carcinoma in a Dog

    OpenAIRE

    Chang-hyun Song1§, Sae-kwang Ku2§, Hwan-soo Jang3, Eun-young Kye, Sung-ho Yun, Kwang-ho Jang and Young-sam Kwon*

    2012-01-01

    A 10-year-old, female, Yorkshire Terrier was presented with a left lower eyelid mass. No other abnormality was detected on affected eye in a general eye examination. The mass was surgically removed and histologically diagnosed as a squamous cell carcinoma. The advancement flap used in this case may be an appropriate therapeutic choice for eyelid squamous cell carcinoma in dogs.

  14. Xenotransplanted human prostate carcinoma (DU145) cells develop into carcinomas and cribriform carcinomas: ultrastructural aspects.

    Science.gov (United States)

    Gilloteaux, Jacques; Jamison, James M; Neal, Deborah R; Summers, Jack L; Taper, Henryk S

    2012-10-01

    Androgen-independent, human prostate carcinoma cells (DU145) develop into solid, carcinomatous xenotransplants on the diaphragm of nu/nu mice. Tumors encompass at least two poorly differentiated cell types: a rapidly dividing, eosinophilic cell comprises the main cell population and a few, but large basophilic cells able to invade the peritoneal stroma, the muscular tissue, lymph vessels. Poor cell contacts, intracytoplasmic lumina, and signet cells are noted. Lysosomal activities are reflected by entoses and programmed cell deaths forming cribriform carcinomas. In large tumors, degraded cells may align with others to facilitate formation of blood supply routes. Malignant cells would spread via ascites and through lymphatics.

  15. CDC27 protein is involved in radiation response in squamous cell cervix carcinoma.

    Science.gov (United States)

    Rajkumar, T; Gopal, G; Selvaluxmi, G; Rajalekshmy, K R

    2005-10-01

    In the present study, an attempt was made to identify genes involved in radiation response in cervix carcinoma. Differential display technique was used to study the expression profiles of tumour biopsy samples obtained from patients, responding and not responding to treatment. The samples were obtained prior to radiotherapy and subsequent to treatment with Tele-radiation at 10 Gray (Gy). One of the differentially expressed cDNAs, when sequenced was identified to be CDC27. Immuno-histochemical analysis of pre- and post-treated tumour samples from fifteen patients showed the downregulation of expression of CDC27 protein in seven patients. Down-regulation was associated with poorer response to radiotherapy. Cervical cancer cell lines SiHa and C33A were irradiated and their nuclei were stained for expression of CDC27 and analyzed using fluorescent-activated cell sorting (FACS). Down-regulation of CDC27 protein in the irradiated SiHa cell line was associated with greater survival fraction, compared to the irradiated C33A cell line, which had only slight fall in the level of CDC27 protein. This is the first study to suggest a role for CDC27 in radiation response. However, a larger cohort is needed to further confirm the value of CDC27 protein as a predictive marker, for radiation response in cervix cancer.

  16. Reynosin protects against neuronal toxicity in dopamine-induced SH-SY5Y cells and 6-hydroxydopamine-lesioned rats as models of Parkinson's disease: Reciprocal up-regulation of E6-AP and down-regulation of α-synuclein.

    Science.gov (United States)

    Ham, Ahrom; Kim, Dong-Woo; Kim, Kyeong Ho; Lee, Sung-Jin; Oh, Ki-Bong; Shin, Jongheon; Mar, Woongchon

    2013-08-01

    Aggregation of α-synuclein (ASYN) is considered a major determinant of neuronal loss in Parkinson's disease (PD). E6-associated protein (E6-AP), an E3 ubiquitin protein ligase, has been known to promote the degradation of α-synuclein. The aim of this study was to assess the effects of the sesquiterpene lactone reynosin on dopamine (DA)-induced neuronal toxicity and regulation of E6-associated protein and α-synuclein proteins in both in vitro and in vivo models of Parkinson's disease. Usi"ng flow cytometry and western blot analysis, we determined that reynosin significantly protected both against cell death from dopamine-induced toxicity in human neuroblastoma SH-SY5Y cells and against the loss of tyrosine hydroxylase (TH)-positive cells in 6-hydroxydopamine (6-OHDA)-lesioned rats (a rodent Parkinson's disease model system). In addition, reynosin made up-regulation of E6-associated protein expression and down-regulation of the over-expression of α-synuclein protein in both dopamine-treated SH-SY5Y cells and 6-hydroxydopamine-lesioned rats. These results suggest that the protective effect of reynosin against dopamine-induced neuronal cell death may be due to the reciprocal up-regulation of E6-associated protein and down-regulation of α-synuclein protein expression.

  17. Growth arrest and apoptosis of human hepatocellular carcinoma cells induced by hexamethylene bisacetamide

    Institute of Scientific and Technical Information of China (English)

    Gao-Liang Ouyang; Qiu-Feng Cai; Min Liu; Rui-Chuan Chen; Zhi Huang; Rui-Sheng Jiang; Fu Chen; Shui-Gen Hong; Shi-Deng Bao

    2004-01-01

    AIM: To investigate the cellular effects of hybrid polar compound hexamethylene bisacetamide (HMBA) on the growth and apoptosis of human hepatocellular carcinoma cells and to provide the molecular mechanism for potential application of HMBA in the treatment of liver cancer.METHODS: Effects of HMBA on the growth of human hepatocellular carcinoma SMMC-7721 cells were assayed by MTT chronometry. Apoptosis induced by HMBA was detected by phase-contrast microscopy, flow cytometry,propidium iodide staining and immunocytochemical analysis.RESULTS: The growth of SMMC-7721 cells was significantly inhibited by HMBA, and the growth inhibitory rate was 51.1%, 62.6%, 68.7% and 73.9% respectively after treatment with 5.0, 7.5, 10.0 and 12.5 mmol/L of HMBA.In the cells treated with 10 mmol/L of HMBA for 72 h, the population of cells at sub-G1 phase significantly increased,and the apoptotic bodies and condensed nuclei were detected. Moreover, treatment of SMMC-7721 cells with 10 mmol/L of HMBA down-regulated the expression of Bcl2 anti-apoptotic protein, while slightly up-regulated the level of pro-apoptotic protein Bax.CONCLUSION: Treatment with 10.0 mmol/L of HMBA can significantly inhibit the growth and induce apoptosis of human hepatocellular carcinoma SMMC-7721 cells by decreasing the ratio of Bcl-2 to Bax.

  18. Treatment Options by Stage (Merkel Cell Carcinoma)

    Science.gov (United States)

    ... Cancer Skin Cancer Screening Research Merkel Cell Carcinoma Treatment (PDQ®)–Patient Version General Information About Merkel Cell ... Certain factors affect prognosis (chance of recovery) and treatment options. The prognosis (chance of recovery ) and treatment ...

  19. Low intensity ultrasound-induced apoptosis in human gastric carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Yi Feng; Zhong-Min Tian; Ming-Xi Wan; Zhao-Bin Zheng

    2008-01-01

    ALIM:To investigate the low intensity ultrasound(US)-induced apoptosis in human gastric carcinoma cells and its potential mechanism and to suggest a new therapeutic approach to gastric carcinoma.METHODS:Human SGC-7901 gastric carcinoma cells were cultured in vitro and irradiated by low intensity US for 10 min at different intensities with different incubation times after irradiation.Morphologic changes were examined under microscope with trypan blue staining and then the percentage of early apoptotic cells was detected by flow cytometry(FCM)with double staining of fluorescein isothiocyanate(FITC)-Annexin V/propidium iodide(PI).Two-dimensional electrophoresis(2DE)was used to get the protein profile and some proteins differently expressed after US irradiation were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF-MS).Functional analysis was performed to investigate the mechanism of US-induced cell apoptosis.RESULTS:The percentage of apoptotic cells increased about 10% after US irradiation(12.0 W/cm2,12 h culture).The percentage of early apoptosis and secondary necrosis in the US-irradiated cells increased with the increased US intensity.Moreover,apoptotic cells increased with the increased culture time after US irradiation and reached its maximum at about 12 h.Several new proteins appeared after US irradiation and were up or down regulated more than 2 times.Some heat shock proteins(HSPs)were found to be associated with the signal process simulating the apoptosis of cells.CONCLUSION:Low intensity US could induce apoptosis in human gastric carcinoma cells.US-induced apoptosis is related to US intensity/culture time.US-induced apoptosis may be caspases-dependent and endoplasmic reticulum(ER)stress-triggered apoptosis may also contribute to it.Proteomic experimental system is useful in finding the protein alteration in carcinoma cells after US irradiation,helping to develop a new cancer therapy.

  20. MicroRNA-24 increases hepatocellular carcinoma cell metastasis and invasion by targeting p53: miR-24 targeted p53.

    Science.gov (United States)

    Chen, Li; Luo, Liang; Chen, Wei; Xu, Hong-Xu; Chen, Fan; Chen, Lian-Zhou; Zeng, Wen-Tao; Chen, Jing-Song; Huang, Xiao-Hui

    2016-12-01

    MicroRNA-24 (miR-24), a member of the miRNA family, functions as an oncogene in various types of human cancer. However, the underlying mechanisms of miR-24 involvement in the development and progression of hepatocellular carcinoma (HCC) remain poorly understood. The present study revealed that miRNA-24 down-regulates p53 through binding to the 3'-UTR of p53 mRNA based on a luciferase reporter assay, and that the expression level of miR-24 could affect the invasion of HCC lines via p53. Down-regulation of p53 significantly attenuated the inhibitory effects of miR-24 knockdown on the invasion of HCC cells, suggesting that miR-24 could be a potential target for HCC treatment. Moreover, our results revealed that miR-24 expression was significantly increased in HCC metastatic tumor tissues compared with matched non-metastatic tumor tissues, and that the up-regulation of miR-24 was significantly associated with down-regulation of p53 in the HCC tissues. In conclusion, this study demonstrates that miR-24 functions as an oncogene in HCC, at least partly by promoting cell invasion through down-regulation of p53. Therefore, miR-24 may be a potential therapeutic target for treatment of HCC.

  1. Identification of genes associated with cisplatin resistance in human oral squamous cell carcinoma cell line

    Directory of Open Access Journals (Sweden)

    Zhang Ping

    2006-09-01

    Full Text Available Abstract Background Cisplatin is widely used for chemotherapy of head and neck squamous cell carcinoma. However, details of the molecular mechanism responsible for cisplatin resistance are still unclear. The aim of this study was to identify the expression of genes related to cisplatin resistance in oral squamous cell carcinoma cells. Methods A cisplatin-resistant cell line, Tca/cisplatin, was established from a cisplatin-sensitive cell line, Tca8113, which was derived from moderately-differentiated tongue squamous cell carcinoma. Global gene expression in this resistant cell line and its sensitive parent cell line was analyzed using Affymetrix HG-U95Av2 microarrays. Candidate genes involved in DNA repair, the MAP pathway and cell cycle regulation were chosen to validate the microarray analysis results. Cell cycle distribution and apoptosis following cisplatin exposure were also investigated. Results Cisplatin resistance in Tca/cisplatin cells was stable for two years in cisplatin-free culture medium. The IC50 for cisplatin in Tca/cisplatin was 6.5-fold higher than that in Tca8113. Microarray analysis identified 38 genes that were up-regulated and 25 that were down-regulated in this cell line. Some were novel candidates, while others are involved in well-characterized mechanisms that could be relevant to cisplatin resistance, such as RECQL for DNA repair and MAP2K6 in the MAP pathway; all the genes were further validated by Real-time PCR. The cell cycle-regulated genes CCND1 and CCND3 were involved in cisplatin resistance; 24-hour exposure to 10 μM cisplatin induced a marked S phase block in Tca/cisplatin cells but not in Tca8113 cells. Conclusion The Tca8113 cell line and its stable drug-resistant variant Tca/cisplatin provided a useful model for identifying candidate genes responsible for the mechanism of cisplatin resistance in oral squamous cell carcinoma. Our data provide a useful basis for screening candidate targets for early diagnosis

  2. 2-Methoxyestradiol induces cell cycle arrest and apoptosis of nasopharyngeal carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Ning-ning ZHOU; Xiao-feng ZHU; Jun-ming ZHOU; Man-zhi LI; Xiao-shi ZHANG; Peng HUANG; Wen-qi JIANG

    2004-01-01

    AIM: To investigate 2-methoxyestradiol induced apoptosis and its mechanism of action in CNE2 cell lines.METHODS: CNE2 cells were cultured in RPMI-1640 medium and treated with 2-methoxyestradiol in different concentrations. MTT assay was used to detect growth inhibition. Flow cytometry and DNA ladders were used to detect apoptosis. Western blotting was used to observe the expression of p53, p21WAF1, Bax, and Bcl-2 protein.RESULTS: 2-methoxyestradiol inhibited proliferation of nasopharyngeal carcinoma CNE2 cells with IC50 value of2.82 μrnol/L. The results of flow cytometry showed an accumulation of CNE2 cells in G2/M phase in response to2-methoxyestradiol. Treatment of CNE2 cells with 2-methoxyestradiol resulted in DNA fragmentation. The expression levels of protein p53 and Bcl-2 decreased following 2-methoxyestradiol treatment in CNE2 cells, whereas Bax and p21WAF1 protein expression were unaffected after treatment with 2-methoxyestradiol. CONCLUSION:These results suggest that 2-methoxyestradiol induced cell cycle arrest at G2/M phase and apoptosis of CNE2 cells which was associated to Bcl-2 down-regulation.

  3. Tubulocystic carcinoma of kidney associated with papillary renal cell carcinoma

    Directory of Open Access Journals (Sweden)

    Mahesh Deshmukh

    2011-01-01

    Full Text Available Tubulocystic renal cell carcinoma (TCRCC is a rare variant of renal cell carcinoma, which has distinct histology but there is some controversy about its association with papillary renal cell carcinoma (PRCC and cell of origin in literature. We report an 18-year-old girl with the rare TCRCC of kidney associated with PRCC with metastases to the para-aortic nodes. The patient presented with hematuria and a right renal mass with enlarged regional nodes for which a radical nephrectomy with retroperitoneal lymph node dissection was done. On gross examination, a solid cystic lesion involving the lower pole and middle pole of the kidney measuring 12x9x9 cm was seen along with an additional cystic lesion in upper pole of kidney. Microscopically the main tumor showed the typical histology of a tubulocystic carcinoma with multiple cysts filled with secretions lined by variably flattened epithelium with hobnailing of cells. The mass in the upper pole was a high-grade PRCC and the nodal metastases had morphology similar to this component. To conclude, at least a small but definite subset of TCRCC is associated with PRCC, and cases associated with PRCC do seem to have a higher propensity for nodal metastasis as in the case we report.

  4. Nevoid Basal Cell Carcinoma Syndrome (Gorlin Syndrome).

    Science.gov (United States)

    Bresler, Scott C; Padwa, Bonnie L; Granter, Scott R

    2016-06-01

    Nevoid basal cell carcinoma syndrome, or basal cell nevus syndrome (Gorlin syndrome), is a rare autosomal dominantly inherited disorder that is characterized by development of basal cell carcinomas from a young age. Other distinguishing clinical features are seen in a majority of patients, and include keratocystic odontogenic tumors (formerly odontogenic keratocysts) as well as dyskeratotic palmar and plantar pitting. A range of skeletal and other developmental abnormalities are also often seen. The disorder is caused by defects in hedgehog signaling which result in constitutive pathway activity and tumor cell proliferation. As sporadic basal cell carcinomas also commonly harbor hedgehog pathway aberrations, therapeutic agents targeting key signaling constituents have been developed and tested against advanced sporadically occurring tumors or syndromic disease, leading in 2013 to FDA approval of the first hedgehog pathway-targeted small molecule, vismodegib. The elucidation of the molecular pathogenesis of nevoid basal cell carcinoma syndrome has resulted in further understanding of the most common human malignancy.

  5. Effects of down-regulatedβ-catenin on epithelial mesenchymal transition, invasion and migration ability of breast cancer cell line MDA-MB-231%β-catenin下调对乳腺癌MDA-MB-231细胞上皮间质转化及侵袭迁移能力的影响

    Institute of Scientific and Technical Information of China (English)

    许光伟; 曹旭晨

    2015-01-01

    Objective To investigate the effect ofβ-catenin on epithelial mesenchymal phenotype and invasion migra⁃tion ability of breast cancer cell line MDA-MB-231. Methods Small interfering RNA (siRNA) targetingβ-catenin plas⁃mid was transfected into MDA-MB-231 cell line. Immunofluorescence staining was used to observe the expression of β-catenin. The expressions of epithelial cell marker E-cadherin and mesenchymal marker vimentin, epithelial mesenchymal transition correlation factor Twist1 and Snail were detected by Western blot assay. Invasion and migration ability was com⁃pared by transwell invasion and wound healing assay between control group, the MDA-MB-231 group andβ-catenin down-regulated group. Results Immunofluorescence staining showed thatβ-catenin was expressed in cell membrane, cytoplasm or nucleus in MDA-MB-231 group and control group. There was a decreased expression in β-catenin down-regulated group, and no expression in cytoplasm or nucleus. The expression of E-cadherin was increased, while vimentin, Twist 1 and Snail expression decreased inβ-catenin down-regulated cells. Transwell invasion and wound healing assay results proved that transmembrane cell number and migration distance were significantly lower in β-catenin down-regulated group than those of MDA-MB-231 group and control group (P<0.05). Conclusion The down-regulation ofβ-catenin inhibits Wnt/β-catenin activation that decreases the mesenchymal phenotype but increases epithelial phenotype of breast cancer cells MDA-MB-231, and which reduces the cell invasion and migration ability in vitro.%目的:探讨β-连环蛋白(β-catenin)对乳腺癌细胞系MDA-MB-231细胞上皮间质表型和侵袭迁移能力的影响。方法用β-catenin siRNA质粒转染乳腺癌MDA-MB-231细胞(对照组),免疫荧光染色观察β-catenin的表达情况。Western blot检测上皮标记蛋白E-cadherin、间质标记蛋白vimentin及上皮间质转化相关调控因子Twist1和Snail的

  6. Deguelin induces both apoptosis and autophagy in cultured head and neck squamous cell carcinoma cells.

    Directory of Open Access Journals (Sweden)

    Yan-li Yang

    Full Text Available Head and neck squamous cell carcinoma (HNSCC represents more than 5% of all cancers diagnosed annually in United States and around the world. Despite advances in the management of patients with this disease, the survival has not been significantly improved, and the search for potential alternative therapies is encouraging. Here we demonstrate that deguelin administration causes a significant HNSCC cell death. Deguelin induces both cell apoptosis and autophagy by modulating multiple signaling pathways in cultured HNSCC cells. Deguelin inhibits Akt signaling, and down-regulates survivin and cyclin-dependent kinase 4 (Cdk4 expressions, by disrupting their association with heat shock protein-90 (Hsp-90. Deguelin induces ceramide production through de novo synthase pathway to promote HNSCC cell death. Importantly, increased ceramide level activates AMP-activated protein kinase (AMPK, which then directly phosphorylates Ulk1 and eventually leads to cell autophagy. We found that a low dose of deguelin sensitized HNSCC cells to 5-FU. Finally, using a nude mice Hep-2 xenograft model, we also showed a significant anti-tumor ability of deguelin in vivo. Together, we suggest that deguelin may represent a novel and effective chemo-agent against HNSCC.

  7. Cisplatin, Radiation Therapy, and Pembrolizumab in Treating Patients With Stage III-IV Head and Neck Squamous Cell Carcinoma

    Science.gov (United States)

    2016-05-16

    Stage III Hypopharyngeal Squamous Cell Carcinoma; Stage III Laryngeal Squamous Cell Carcinoma; Stage III Oral Cavity Squamous Cell Carcinoma; Stage III Oropharyngeal Squamous Cell Carcinoma; Stage IVA Hypopharyngeal Squamous Cell Carcinoma; Stage IVA Laryngeal Squamous Cell Carcinoma; Stage IVA Oral Cavity Squamous Cell Carcinoma; Stage IVA Oropharyngeal Squamous Cell Carcinoma; Stage IVB Hypopharyngeal Squamous Cell Carcinoma; Stage IVB Laryngeal Squamous Cell Carcinoma; Stage IVB Oral Cavity Squamous Cell Carcinoma; Stage IVB Oropharyngeal Squamous Cell Carcinoma

  8. Neglected giant scalp Basal cell carcinoma

    DEFF Research Database (Denmark)

    Larsen, Anne Kristine; El-Charnoubi, Waseem-Asim Ghulam; Gehl, Julie;

    2014-01-01

    SUMMARY: Rarely, basal cell carcinoma grows to a giant size, invading the underlying deep tissue and complicating the treatment and reconstruction modalities. A giant basal cell carcinoma on the scalp is in some cases treated with a combination of surgery and radiation therapy, resulting in local...... control, a satisfactory long-term cosmetic and functional result. We present a case with a neglected basal cell scalp carcinoma, treated with wide excision and postoperative radiotherapy, reconstructed with a free latissimus dorsi flap. The cosmetic result is acceptable and there is no sign of recurrence...

  9. ACANTHOLYTIC SQUAMOUS CELL CARCINOMA OF PREPUCE

    Directory of Open Access Journals (Sweden)

    Mamina

    2014-03-01

    Full Text Available An uncircumcised 65 year male, with history of phimosis presented with retention of urine and ulceration and bleeding in the prepuce. Circumcision was done under local anesthesia which revealed an ulcero-proliferative growth involving the prepuce and glans. The prepucial skin was sent for histopathological examination. The diagnosis was histopathologically confirmed as Acantholytic Squamous Cell Carcinoma. Acantholytic squamous cell carcinoma is a highly malignant, unusual variant of squamous cell carcinoma invading deeper anatomic structures and is associated with a higher incidence of regional metastasis and mortality.

  10. Neglected Giant Scalp Basal Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Anne Kristine Larsen, MD

    2014-03-01

    Full Text Available Summary: Rarely, basal cell carcinoma grows to a giant size, invading the underlying deep tissue and complicating the treatment and reconstruction modalities. A giant basal cell carcinoma on the scalp is in some cases treated with a combination of surgery and radiation therapy, resulting in local control, a satisfactory long-term cosmetic and functional result. We present a case with a neglected basal cell scalp carcinoma, treated with wide excision and postoperative radiotherapy, reconstructed with a free latissimus dorsi flap. The cosmetic result is acceptable and there is no sign of recurrence 1 year postoperatively.

  11. Uptake of donor lymphocytes treated with 8-methoxypsoralen and ultraviolet A light by recipient dendritic cells induces CD4{sup +}CD25{sup +}Foxp3{sup +} regulatory T cells and down-regulates cardiac allograft rejection

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, De-Hua [Organ Transplant Center, Chinese PLA 309th Hospital, No. 17A Hei-Shan-Hu Road, Beijing 100091 (China); Dou, Li-Ping [Department of Hematology, Chinese PLA General Hospital, No. 28 Fu-Xing Road, Beijing 100853 (China); Wei, Yu-Xiang; Du, Guo-Sheng; Zou, Yi-Ping; Song, Ji-Yong; Zhu, Zhi-Dong; Cai, Ming; Qian, Ye-Yong [Organ Transplant Center, Chinese PLA 309th Hospital, No. 17A Hei-Shan-Hu Road, Beijing 100091 (China); Shi, Bing-Yi, E-mail: shibingyi@medmail.com.cn [Organ Transplant Center, Chinese PLA 309th Hospital, No. 17A Hei-Shan-Hu Road, Beijing 100091 (China)

    2010-05-14

    Extracorporeal photopheresis (ECP) is an effective immunomodulatory therapy and has been demonstrated to be beneficial for graft-vs-host disease and solid-organ allograft rejection. ECP involves reinfusion of a patient's autologous peripheral blood leukocytes treated ex vivo with 8-methoxypsoralen and UVA light radiation (PUVA). Previous studies focused only on ECP treatment of recipient immune cells. Our study is the first to extend the target of ECP treatment to donor immune cells. The results of in vitro co-culture experiments demonstrate uptake of donor PUVA-treated splenic lymphocytes (PUVA-SPs) by recipient immature dendritic cells (DCs). Phagocytosis of donor PUVA-SPs does not stimulate phenotype maturation of recipient DCs. In the same co-culture system, donor PUVA-SPs enhanced production of interleukin-10 and interferon-{gamma} by recipient DCs and impaired the subsequent capability of recipient DCs to stimulate recipient naive T cells. Phagocytosis of donor PUVA-SP (PUVA-SP DCs) by recipient DCs shifted T-cell responses in favor of T helper 2 cells. Infusion of PUVA-SP DCs inhibited cardiac allograft rejection in an antigen-specific manner and induced CD4{sup +}CD25{sup high}Foxp3{sup +} regulatory T cells. In conclusion, PUVA-SP DCs simultaneously deliver the donor antigen and the regulatory signal to the transplant recipient, and thus can be used to develop a novel DC vaccine for negative immune regulation and immune tolerance induction.

  12. Down-Regulation of miR-3928 Promoted Osteosarcoma Growth

    Directory of Open Access Journals (Sweden)

    Haidong Xu

    2014-05-01

    Full Text Available Background: Osteosarcoma is the most common primary bone malignancy in children and young adults. Most failures of osteosarcoma treatment were due to resistance to chemotherapy. Development of new therapy required elucidation underlying molecular mechanism. Many miRNAs have been proved to be