Research ArticleExperimental Studies

Phenethyl Isothiocyanate Inhibits Migration and Invasion of Human Gastric Cancer AGS Cells through Suppressing MAPK and NF-κB Signal Pathways

MEI-DUE YANG, KUANG-CHI LAI, TUNG-YUAN LAI, SHU-CHUN HSU, CHAO-LIN KUO, CHUN-SHU YU, MENG-LIANG LIN, JAI-SING YANG, HSIU-MAAN KUO, SHIN-HWAR WU and JING-GUNG CHUNG
Anticancer Research June 2010, 30 (6) 2135-2143;

Abstract

Cell motility involves metastasis suppressors and other regulators that play an important role in tumor invasion and metastasis. Phenethyl isothiocyanate (PEITC), found in dietary cruciferous vegetables, has been found to exhibit antitumor properties and therefore is of special interest for the development of chemopreventive and chemotherapeutic agent for human cancers. Here, we report that in addition to its function as an anticancer agent, and PEITC can inhibit migration and invasion through the extracellular signal-regulated kinases 1/2 (ERK1/2), protein kinase C (PKC) and nuclear factor-kappaB (NF-κB) signaling pathways in human gastric cells. The results from wound healing and Boyden chamber assays (migration and invasion) assay indicated that PEITC exhibited an inhibitory effect on the migration and invasion of AGS cells. Results from Western blotting examination demonstrated that PEITC exerted an inhibitory effect on the ERK1/2, mitogen-activated protein kinase kinase 7 (MKK7), MAP kinase kinase kinase 3 (MEKK3), son of sevenless 1 (SOS1), PKC, Ras homolog gene family, member A (Rho A) and urokinase-type plasminogen activator (uPA), causing the inhibition of matrix metallopeptidase-2 (MMP-2) and -9 then followed by the inhibition of invasion and migration of GAS cells in vitro. PEITC also inhibited Ras, growth factor receptor-bound protein 2 (GRB2), vascular endothelial growth factor (VEGF), focal adhesion kinase (FAK), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), causing inhibition of cell proliferation of AGS cells. Results from real-time PCR showed that PEITC inhibited the gene expressions of MMP-2, -7 and -9, FAK and RhoA after PEITC treatment for 24 and 48 h of AGS cells. Taken together, these findings may provide insight into a new mechanisms and functions of PEITC in migration and invasion of human gastric cancer AGS cells. Our data imply that molecular targeting of PKC leading to the inhibition of MMP-2 and -9 might be a useful strategy for the inhibition of migration and invasion of human gastric cancer.

Stomach cancer is the fourth most common cancer and the second most common cause of cancer death in the world (1). In Taiwan, about 10 people per 100 thousand die annually from stomach cancer, based on reports from Department of Health, R.O.C. (Taiwan) in year 2008. Although many treatments including surgery, radiation, chemotherapy, or combination of radiotherapy with chemotherapy have been used for patients, the mortality of stomach cancer remains high. It was reported that dietary intake of cruciferous vegetables may reduce the risk of various types of malignancies and organic isothiocyanates are important components of such vegetables (2, 3). Phenethyl isothiocyanate (PEITC) has been shown to possess chemopreventive activity (4). PEITC inhibits cytochrome P450 (CYP) enzymes and induces phase II detoxification enzymes and apoptosis in HT-29 cells (5). In an animal model, PEITC inhibited 4-(methylnitrosamino)-1-(3-pyridyl)-1-butone-induced pulmonary neoplasia (6, 7) and also inhibited azoxymethane-induced colonic aberrant crypt foci formation (8).

Metastasis of cancer cells is associated with various steps and cytophysiological changes such as changed adhesion between cells and the extracellular matrix (ECM) and this disrupted intercellular interaction and the ECM degradation are associated with tumor invasion and migration. Overexpression of matrix metalloproteinases (MMPs) and urokinase-type plasminogen activator (uPA) are associated with the metastasis of cancer cells. Much evidence has shown that MMPs are involved in the invasion and metastasis of various tumor cells (9-11). It was reported that MMP-2 and MMP-9 are capable of degrading most ECM components that form the basal membrane (12-14). Several studies demonstrated that inhibition of MMP expression or enzyme activity can be used as targets for preventing cancer metastasis (15-16).

Many experiments have demonstrated that PEITC can induce apoptosis in many types of human cancer cells (17-21), but there is no information to show whether PEITC inhibits the migration and invasion of stomach cancer cells. Therefore, in the present study, we investigated the effect of PEITC on the migration and invasion of human stomach cancer AGS cells in vitro.

Materials and Methods

Chemicals and reagents. PEITC, dimethyl sulfoxide (DMSO), potassium phosphates, propidium iodide and trypan blue were purchased from Sigma Chemical Co. (St. Louis, MO, USA). RPMI-1640 medium, fetal bovine serum (FBS), L-glutamine, penicillin-streptomycin and trypsin-EDTA were obtained from Gibco (Invitrogen, Carlsbad, CA, USA). Primary antibodies used for Western blotting analysis were obtained as follows: antibodies for MMP-2, MMP-9, inductible nitric oxide synthase (iNOS), nuclear factor-kappaB (NF-κB) p65, cyclooxygenase-2 (COX-2), uPA, mitogen-activated protein kinase kinase 7 (MKK7), MAP kinase kinase kinase 3 (MEKK3), focal adhesion kinase (FAK), extracellular signal-regulated kinases 1/2 (ERK1/2), c-Jun NH2-terminal kinases 1/2 (JNK1/2), p-p38, Ras, growth factor receptor-bound protein 2 (GRB2), vascular endothelial growth factor (VEGF), Ras homolog gene family, member A (Rho A), Rho-associated coiled coil-containing protein kinases 1 (ROCK1), son of sevenless 1 (SOS1), phosphatidylinositol 3-kinase (PI3K), protein kinase C (PKC) and β-Actin were purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA) and diluted before use.

AGS cell line. Human gastric cancer AGS cells were obtained from the Food Industry Research and Development Institute (Hsinchu, Taiwan, R.O.C.). Cells were cultured with 88% RPMI-1640 medium with 2 mM L-glutamine and supplemented with 10% FBS, and 1% penicillin-streptomycin (100 Units/ml penicillin and 100 μg/ml streptomycin) and were incubated in a humidified atmosphere of 5% CO2 and 95% air at 37°C.

Wound-healing assay. AGS cell migration was examined by using a wound-healing assay. In brief, AGS cells (1×105 cells/well) were placed for 24 h in six-well plates and at confluence a wound was made with a pipette tip followed by washing with serum-free medium to remove cell debris. They were then photographed under phase-contrast microscopy (time=0) and then incubated in media with or without PEITC (0, 0.01 and 0.25 μM) at 37°C and 5% CO2 and allowed to migrate into the wound area for up to 24 h at 37°C. Then cells were gently washed with phosphate buffered saline (PBS) and the wound area photographed under phase-contrast microscope (22). Experiments were performed in triplicate.

In vitro migration and invasion assays. The migration of AGS cells was also measured by chemotactic directional migration by using a 24-well Transwell insert. The 8 μm pore filters (Millipore, MA, USA) were coated with 30 μg type I collagen (Millipore) for 1 h and AGS cells (104 cells/0.4 ml RPMI-1640 medium) were placed in the upper chamber with or without PEITC (0.25 or 0.50 μM) and allowed to undergo migration for 24 and 48 h. The non-migrated cells in the upper chamber were removed with a cotton swab. The filters were stained with 2% crystal violet. Migrated cells adherent to the underside of the filter were counted and photographed under a light microscope at ×200 (23-24).

The invasion of AGS cells was measured using Matrigel-coated transwell cell culture chambers (8 μm pore size) as previously described (23, 25). After cells were cultured for 24 h in serum-free RPMI-1640 medium, they were collected and resuspended in serum-free medium, and then were placed in the upper chamber of the transwell insert (5×104 cells/well) and treated with 0.5% DMSO (as a control) or PEITC (0.25 or 0.50 μM). RPMI-1640 medium containing 10% FBS was placed in the lower chamber. All cells in each treatment were incubated for 24 or 48 h at 37°C in a humidified atmosphere with 95% air and 5% CO2. The non-invasive cells maintained in the upper chamber were removed by wiping with a cotton swab and the invasive cells were fixed with 4% formaldehyde in PBS and stained with 2% crystal violet in 2% ethanol. The cells (invasive cells) in the lower surface of the filter which penetrated through the Matrigel were counted and photographed under a light microscope at ×200 (23). Each treatment, including control, was assayed twice and three independent experiments were performed.

Western blotting analysis. AGS cells (1×106 cells/well) were cultured in 6-well plates and grown for 24 h. PEITC was added to cells in each wells at a final concentration of 2.5 μM, while DMSO (solvent) alone was added to control cells. All cells in the wells were incubated at 37°C for 0, 6, 12, 24 and 48 h. The cells were then collected and resuspended in ice-cold 50 mM potassium phosphate buffer (pH 7.4) containing 2 mM EDTA and 0.1% Triton X-100 for sonication and centrifugation at 13,000 g for 10 min at 4°C to remove cell debris. The supernatant was collected and total protein concentration of each sample was determined using a Bio-Rad protein assay kit (Hercules, CA, USA) with bovine serum albumin (BSA) as the standard. SDS gel electrophoresis and Western blotting were conducted to determine the effects of PEITC on protein levels of MMP-2, MMP-9, iNOS, NF-κB p65, COX-2, uPA, MKK7, MEKK3, FAK, ERK1/2, JNK1/2 and p-p38, Ras, GRB2, VEGF, RhoA, ROCK1, SOS1, PI3K, PKC and β-actin as described previously (26-27).

Real-time PCR of MMP-2, -7, and -9, FAK and Rho A. AGS cells (1×106 cells/well) were cultured in 6-well plates and grown for 24 h. PEITC was added to cells in each well for a final concentration of 2.5 μM for 24 and 48 h. Cells were collected and total RNA was extracted using Qiagen RNeasy Mini Kit as described previously (25, 28). RNA samples were reverse-transcribed at 42°C with High Capacity cDNA Reverse Transcription Kit for 30 min according to the protocol of the supplier (Applied Biosystems, Foster City, CA, USA). Quantitative PCR conditions were: 2 min at 50°C, 10 min at 95°C, and 40 cycles of 15 s at 95°C; 1 min at 60°C using 1 μl of the cDNA reverse-transcribed as described above, 2X SYBR Green PCR Master Mix (Applied Biosystems) and 200 nM of forward and reverse primers as shown in Table I. Applied Biosystems 7300 Real-Time PCR system was used for each assay in triplicate and expression fold-changes were derived using the comparative CT method.

Statistical analysis. Statistical differences were evaluated using Student's t-test and considered significant at p<0.05. All data are presented as means±S.D. of three experiments.

Results

Effects of PEITC on migration of AGS cells. To determine the effect of PEITC on cell migration, AGS cells were incubated with different concentrations of PEITC for 24 h, then wound healing assay was carried out; the results are shown in Figure 1. An apparent and gradual increase of cells in the denuded zone was observed by light microscopy. The quantitative data in Figure 1B indicate a significant inhibition of cell migration and this was also time-dependent.

Effect of PEITC on migration and invasion of AGS cells. To further examine the effect of PEITC on cell migration, AGS cells were incubated with different concentrations of PEITC for 24 and 48 h in a migration assay. Results from the migration assay are shown in Figure 2A and B, indicating that PEITC had a significant inhibitory effect on cell migration at concentrations between 0.25-0.50 μM and the percentage inhibition was at 44-45% and 38-47% when cells were incubated with PEITC for 24 and 48 h, respectively. AGS cells were also tented for their ability to invade through a filter coated with Matrigel on treatment with PEITC for 24 and 48 h. Significant inhibition of cell invasion occurred in a dose-dependent manner, as shown in Figure 2C and D. The percentage inhibition at 0.25 μM PEITC was 53-55% for 24 and 48 h treatments and at 0.50 μM PEITC for 24 and 48 h treatment, the inhibition ranged from 37-54% (Figure 2D).

Effect of PEITC on levels of proteins associated with migration and invasion in AGS cells. In order to determine the effects of PEITC on the protein levels associated with migration and invasion in AGS cells, Western blotting was applied after the cells were exposed to PEITC. The results are shown in Figure 3 and indicate that PEITC reduced levels of MMP-2 and MMP-9 (Figure 3A) iNOS, COX-2 and uPA (Figure 3B), MKK7, MEKK3, FAK, ERK1/2, JNK1/2 (Figure 3C), Ras, GRB2, VEGF, RhoA, ROCK1, SOS1 and PKC (Figure 3D) but increased protein levels of NF-κB p65 (Figure 3B), p-p38 (Figure 3C) and PI3K (Figure 3D) in the examined AGS cells. These effects may be responsible for the inhibition of migration and invasion of AGS cells.

View this table:
Table I.

Primers sequence used for real-time PCR.

Effects of PEITC on MMP-2, MMP-7, MMP-9, FAK and RhoA mRNA expressions in AGS cells. To investigate whether or not PEITC affected migration- and invasion-associated gene expression in AGS cells, cells were treated with PEITC (2.5 μM) for 0, 24 and 48 h. Total RNA was isolated and gene expressions were determined by real-time PCR. The results are shown in Figure 4. The expression levels of MMP-2, MMP-7, MMP-9, FAK and RhoA were inhibited during PEITC treatment for 24 and 48 h and these effects were time-dependent (Figure 4A and B). The greatest inhibition occurred for RhoA expression (Figure 4B).

Discussion

Cancer cell metastasis involves tumor cell adhesion to the ECM, proteolytic cleavage or destruction of the ECM, and cell migration through the resultant defect. In the present study, we demonstrate that PEITC reduced invasion and migration of AGS cells. The breakdown of biological barriers such as the basement membrane which requires activation of proteolytic enzymes is a critical step for invasion and metastasis (29, 30). Of these basement membrane-degrading enzymes, the MMPs play an important role in tumor angiogenesis, metastasis and growth factor release from the ECM (26). MMP-2 (72-kDa gelatinase A) and MMP-9 (92-kDa gelatinase B) are associated with the invasive metastatic potential of tumor cells (31). It was reported that inhibition of MMP expression or enzyme activity can be used as targets for preventing cancer metastasis (15-16). Thus, the present of cell migration provides many molecular targets for the development of therapeutic agents to inhibit cancer metastasis.

Figure 1.
Figure 1.

Effects of PEITC on the migration of AGS cells: Wound-healing migration assay. AGS cells were placed on the plate and a wound line was made with a tip. The cells were incubated with or without 0.25 and 0.50 μM of PEITC for 0, 24 and 48 h. A: Cell migration was assessed by microscopy at the indicated time points (×100). B: The mean number cells in the denuded zone were shown when compared with control group at 0 h. Each bar represents the mean±S.D. (n=3). *Significantly the different from the control (0 μM) at p<0.05.

Figure 2.
Figure 2.

Effect of PEITC on migration and invasion of AGS cells. AGS cells were treated with PEITC for 24 and 48 h. Cell migration was measured in a Boyden chamber assay with A:polycarbonate filters (pore size, 8 μm); and C: polycarbonate filters were precoated with Matrigel. Migration (B) and invasion (D) ability of AGS cells were quantified by counting the numberof cells that invaded the underside of the porous polycarbonate membrane under microscopy and represent the average of three experiments. *p<0.05 compared with the untreated control(dose 0 μM).

Figure 3.
Figure 3.

Effect of PEITC on the protein levels of proteins associated with migration and invasion in AGS cells. AGS cells were treated with 2.5 μM of PEITC for 6, 12, 24 and 48 h. The total proteins were collected from each sample and the protein levels were measured by SDS-PAGE and Western blotting as described in the Materials and Methods.

Our results showed that PEITC inhibited the migration and invasion of AGS cells through the examinations of wound healing assay and migration and invasion assays and these effects occurred in dose- and time-dependent manners. Furthermore, we found that PEITC reduced the level of proteins associated with migration and invasion, such as MMP-2 and MMP-9, iNOS, COX-2, uPA, MKK7, MEKK3, FAK, ERK1/2, JNK, Ras, GRB2, VEGF, RhoA and ROCK1, but increased protein levels of NF-κB p65, p-p38 and PI3K. Overexpression of MMPs such as MMP-2 (32, 33), MMP-7 (34, 35) and MMP-9 (36) has been observed in gastric cancer. But our result did not show such as effect of PEITC on MMP-7 protein level.

Figure 4.
Figure 4.

Effect of PEITC on MMP-2, MMP-7, MMP-9, FAK and RhoA mRNA expressions in AGS cells. The total RNA was extracted from cells treated with PEITC (2.5 μM) for 0, 24 and 48 h, and RNA samples were reverse-transcribed to obtain cDNA then for real-time PCR as described in the Materials and Methods. The ratios of gene expressions related to that for GAPDH are presented in A and B. Data represents the mean±S.D. of three experiments. *p<0.05 compared with the untreated control.

However, our results also showed that PEITC suppressed MMP-2, MMP-7, MMP-9, FAK and RhoA gene expressions, which may suppress the PKC/MAPK and PI3K/AKT/NF-κB cascades with consequent suppression of migration and invasion by human gastric cancer AGS cells.

Highly metastatic tumors have been shown to overexpression MMP-2, and MMP-9 can be stimulated by TNF-α (37) or growth factors such as VEGF, Epidermal growth factor (EGF) and transforming growth factor beta (TGF-β) (38-39), or Ras oncogene (5, 40) through activation of different intracellular signaling pathways. Results from the present study indicate that PEITC inhibited VEGF and Ras, which may have led to the inhibition of MMP-7 expression. Other investigators also showed that the activation of PKC led to translocation of the protein to membranes and led to control of the expression of MMP-9 through modulating the activation of transcription factors including NF-κB or SP-1 through MAPK and PI3K signaling pathways (40-42). Our results also showed that PEITC promoted the levels of NF-κB in AGS cells.

Figure 5.
Figure 5.

The proposed mechanisms of PEITC inhibition of migration and invasion of human gastric cancer AGS cells.

In the present study, we observed that PEITC inhibited not only phosphorylation of JNK but also VEGF, MMP-9 and uPA expression, suggesting that inhibition of JNK by PEITC is involved in the inhibition of VEGF, MMP-9 and uPA expression. These results indicate that inhibition of JNK activity contributes to the decrease in migration of PEITC-treated AGS cells. The results in the present study also showed that the regulation of NF-κB, and downstream of the PI3K/AKT and MAPK (ERK1/2, p38 and JNK) pathways might be involved in PEITC suppression of MMP-9 expression and invasion in AGS cells. The activation of NF-κB is involved in the induction of the MMP-9 gene associated with the invasion and metastasis of tumor cells (37, 43). We also found that PEITC reduced the JNK and PKC levels (Figure 4). It was reported that resveratrol suppresses MMP-9 expression in phorbol myristate acetate (PMA)-induced human Caski cells by blocking JNK and PKCδ signal transduction (44).

Our results also showed that PEITC reduced the level of uPA, which has been shown to play a major role in the decomposition of basement membranes. The activation of the uPA/uPAR/plasmin proteolytic network is involved in tumor invasion and dissemination of various malignancies (45-46). Thus, the presence of uPA in tumor tissues is a potential prognostic factor, and the levels of uPA and uPAR expression serve as prognostic markers in various malignancies. Several reports indicated that ERK1/2, p38 and JNK play a central role in regulating the expression of MMPs and uPA (47-49). Thus, inhibition of the MAPK pathway might have the potential to prevent invasion, and metastasis for a wide range of tumor types. Other signal pathways, such as the PI3K/AKT signal transduction pathway regulating cell metastasis of prostate cancer PC-3 cells, are closely associated with the development and progress of various tumors (49). In the present study, PEITC also reduced JNK1/2. Future studies are needed to explore the relationship between the expression of MMPs and phosphorylation of JNK in gastric cancer cells treated with PEITC.

In summary, the molecular mechanism of PEITC appears to be via down-regulation of PKC and then blocking of MAPK signaling pathways through NF-κB, as well as uPA, which then led to the inhibition of MMP-2 and MMP-9 (Figure 5). The present results may be relevant to the therapeutic targeting of invasion and migration of gastric cancer cells.

Acknowledgements

This work was supported by Grant CMU-97-276 from China Medical University, Taichung, Taiwan, R.O.C.

  • Received December 18, 2009.
  • Revision received April 20, 2010.
  • Accepted April 28, 2010.

References

  1. ↵
    1. Ajani JA
    : Evolving chemotherapy for advanced gastric cancer. Oncologist 10(Suppl 3): 49-58, 2005.
    OpenUrlAbstract/FREE Full Text
  2. ↵
    1. Bonnesen C,
    2. Eggleston IM,
    3. Hayes JD
    : Dietary indoles and isothiocyanates that are generated from cruciferous vegetables can both stimulate apoptosis and confer protection against DNA damage in human colon cell lines. Cancer Res 61: 6120-6130, 2001.
    OpenUrlAbstract/FREE Full Text
  3. ↵
    1. Nastruzzi C,
    2. Cortesi R,
    3. Esposito E,
    4. Menegatti E,
    5. Leoni O,
    6. Iori R,
    7. Palmieri S
    : In vitro antiproliferative activity of isothiocyanates and nitriles generated by myrosinase-mediated hydrolysis of glucosinolates from seeds of cruciferous vegetables. J Agric Food Chem 48: 3572-3575, 2000.
    OpenUrlCrossRefPubMed
  4. ↵
    1. Stoner GD,
    2. Morrissey DT,
    3. Heur YH,
    4. Daniel EM,
    5. Galati AJ,
    6. Wagner SA
    : Inhibitory effects of phenethyl isothiocyanate on N-nitrosobenzylmethylamine carcinogenesis in the rat esophagus. Cancer Res 51: 2063-2068, 1991.
    OpenUrlAbstract/FREE Full Text
  5. ↵
    1. Zhang Y
    : Cancer-preventive isothiocyanates: measurement of human exposure and mechanism of action. Mutat Res 555: 173-190, 2004.
    OpenUrlCrossRefPubMed
  6. ↵
    1. Wu X,
    2. Kassie F,
    3. Mersch-Sundermann V
    : Induction of apoptosis in tumor cells by naturally occurring sulfur-containing compounds. Mutat Res 589: 81-102, 2005.
    OpenUrlCrossRefPubMed
  7. ↵
    1. Morse MA,
    2. Amin SG,
    3. Hecht SS,
    4. Chung FL
    : Effects of aromatic isothiocyanates on tumorigenicity, O6-methylguanine formation, and metabolism of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in A/J mouse lung. Cancer Res 49: 2894-2897, 1989.
    OpenUrlAbstract/FREE Full Text
  8. ↵
    1. Zhang Y,
    2. Kensler TW,
    3. Cho CG,
    4. Posner GH,
    5. Talalay P
    : Anticarcinogenic activities of sulforaphane and structurally related synthetic norbornyl isothiocyanates. Proc Natl Acad Sci USA 91: 3147-3150, 1994.
    OpenUrlAbstract/FREE Full Text
  9. ↵
    1. Kilian M,
    2. Gregor JI,
    3. Heukamp I,
    4. Hanel M,
    5. Ahlgrimm M,
    6. Schimke I,
    7. Kristiansen G,
    8. Ommer A,
    9. Walz MK,
    10. Jacobi CA,
    11. Wenger FA
    : Matrix metalloproteinase inhibitor RO 28-2653 decreases liver metastasis by reduction of MMP-2 and MMP-9 concentration in BOP-induced ductal pancreatic cancer in Syrian hamsters: inhibition of matrix metalloproteinases in pancreatic cancer. Prostaglandins Leukot Essent Fatty Acids 75: 429-434, 2006.
    OpenUrlCrossRefPubMed
    1. Mizutani K,
    2. Kofuji K,
    3. Shirouzu K
    : The significance of MMP-1 and MMP-2 in peritoneal disseminated metastasis of gastric cancer. Surg Today 30: 614-621, 2000.
    OpenUrlCrossRefPubMed
  11. ↵
    1. Gullu IH,
    2. Kurdoglu M,
    3. Akalin I
    : The relation of gelatinase (MMP-2 and -9) expression with distant site metastasis and tumour aggressiveness in colorectal cancer. Br J Cancer 82: 249, 2000.
    OpenUrlCrossRefPubMed
  12. ↵
    1. Bernhard EJ,
    2. Gruber SB,
    3. Muschel RJ
    : Direct evidence linking expression of matrix metalloproteinase 9 (92-kDa gelatinase/collagenase) to the metastatic phenotype in transformed rat embryo cells. Proc Natl Acad Sci USA 91: 4293-4297, 1994.
    OpenUrlAbstract/FREE Full Text
    1. Donadio AC,
    2. Remedi MM,
    3. Susperreguy S,
    4. Frede S,
    5. Gilardoni MB,
    6. Tang Y,
    7. Pellizas CG,
    8. Yan L
    : Extracellular matrix metalloproteinase inducer (EMMPRIN) and matrix metalloproteinases (MMPs) as regulators of tumor-host interaction in a spontaneous metastasis model in rats. Histochem Cell Biol 130: 1155-1164, 2008.
    OpenUrlCrossRefPubMed
  14. ↵
    1. Cortes-Reynosa P,
    2. Robledo T,
    3. Macias-Silva M,
    4. Wu SV,
    5. Salazar EP
    : Src kinase regulates metalloproteinase-9 secretion induced by type IV collagen in MCF-7 human breast cancer cells. Matrix Biol 27: 220-231, 2008.
    OpenUrlCrossRefPubMed
  15. ↵
    1. Waas ET,
    2. Wobbes T,
    3. Lomme RM,
    4. DeGroot J,
    5. Ruers T,
    6. Hendriks T
    : Matrix metalloproteinase -2 and -9 activity in patients with colorectal cancer liver metastasis. Br J Surg 90: 1556-1564, 2003.
    OpenUrlCrossRefPubMed
  16. ↵
    1. Okada N,
    2. Ishida H,
    3. Murata N,
    4. Hashimoto D,
    5. Seyama Y,
    6. Kubota S
    : Matrix metalloproteinase-2 and -9 in bile as a marker of liver metastasis in colorectal cancer. Biochem Biophys Res Commun 288: 212-216, 2001.
    OpenUrlCrossRefPubMed
  17. ↵
    1. Lee JW,
    2. Cho MK
    : Phenethyl isothiocyanate induced apoptosis via down-regulation of Bcl-2/XIAP and triggering of the mitochondrial pathway in MCF-7 cells. Arch Pharm Res 31: 1604-1612, 2008.
    OpenUrlCrossRefPubMed
    1. Cuddihy SL,
    2. Brown KK,
    3. Thomson SJ,
    4. Hampton MB
    : Induction of apoptosis by phenethyl isothiocyanate in cells overexpressing Bcl-XL. Cancer Lett 271: 215-221, 2008.
    OpenUrlCrossRefPubMed
    1. Thomson SJ,
    2. Brown KK,
    3. Pullar JM,
    4. Hampton MB
    : Phenethyl isothiocyanate triggers apoptosis in Jurkat cells made resistant by the overexpression of Bcl-2. Cancer Res 66: 6772-6777, 2006.
    OpenUrlAbstract/FREE Full Text
    1. Satyan KS,
    2. Swamy N,
    3. Dizon DS,
    4. Singh R,
    5. Granai CO,
    6. Brard L
    : Phenethyl isothiocyanate (PEITC) inhibits growth of ovarian cancer cells by inducing apoptosis: role of caspase and MAPK activation. Gynecol Oncol 103: 261-270, 2006.
    OpenUrlCrossRefPubMed
  21. ↵
    1. Xiao D,
    2. Zeng Y,
    3. Choi S,
    4. Lew KL,
    5. Nelson JB,
    6. Singh SV
    : Caspase-dependent apoptosis induction by phenethyl isothiocyanate, a cruciferous vegetable-derived cancer chemopreventive agent, is mediated by Bak and Bax. Clin Cancer Res 11: 2670-2679, 2005.
    OpenUrlAbstract/FREE Full Text
  22. ↵
    1. Ho YT,
    2. Yang JS,
    3. Li TC,
    4. Lin JJ,
    5. Lin JG,
    6. Lai KC,
    7. Ma CY,
    8. Wood WG,
    9. Chung JG
    : Berberine suppresses in vitro migration and invasion of human SCC-4 tongue squamous cancer cells through the inhibitions of FAK, IKK, NF-kappaB, uPA and MMP-2 and -9. Cancer Lett 279: 155-162, 2009.
    OpenUrlCrossRefPubMed
  23. ↵
    1. Lai KC,
    2. Huang AC,
    3. Hsu SC,
    4. Kuo CL,
    5. Yang JS,
    6. Wu SH,
    7. Chung JG
    : Benzyl isothiocyanate (BITC) inhibits migration and invasion of human colon cancer HT29 cells by inhibiting matrix metalloproteinase-2/-9 and urokinase plasminogen (uPA) through PKC and MAPK signaling pathway. J Agric Food Chem, 2010.
  24. ↵
    1. Lin CC,
    2. Chen JT,
    3. Yang JS,
    4. Lu HF,
    5. Hsu SC,
    6. Tan TW,
    7. Lin YT,
    8. Ma YS,
    9. Ip SW,
    10. Wu JJ,
    11. Li YC,
    12. Chung JG
    : Danthron inhibits the migration and invasion of human brain glioblastoma multiforme cells through the inhibition of mRNA expression of focal adhesion kinase, Rho kinases-1 and metalloproteinase-9. Oncol Rep 22: 1033-1037, 2009.
    OpenUrlPubMed
  25. ↵
    1. Lin HJ,
    2. Su CC,
    3. Lu HF,
    4. Yang JS,
    5. Hsu SC,
    6. Ip SW,
    7. Wu JJ,
    8. Li YC,
    9. Ho CC,
    10. Wu CC,
    11. Chung JG
    : Curcumin blocks migration and invasion of mouse-rat hybrid retina ganglion cells (N18) through the inhibition of MMP-2, -9, FAK, Rho A and Rock-1 gene expression. Oncol Rep 23: 665-670, 2010.
    OpenUrlPubMed
  26. ↵
    1. Lin SY,
    2. Lai WW,
    3. Ho CC,
    4. Yu FS,
    5. Chen GW,
    6. Yang JS,
    7. Liu KC,
    8. Lin ML,
    9. Wu PP,
    10. Fan MJ,
    11. Chung JG
    : Emodin induces apoptosis of human tongue squamous cancer SCC-4 cells through reactive oxygen species and mitochondria-dependent pathways. Anticancer Res 29: 327-336, 2009.
    OpenUrlAbstract/FREE Full Text
  27. ↵
    1. Lai WW,
    2. Yang JS,
    3. Lai KC,
    4. Kuo CL,
    5. Hsu CK,
    6. Wang CK,
    7. Chang CY,
    8. Lin JJ,
    9. Tang NY,
    10. Chen PY,
    11. Huang WW,
    12. Chung JG
    : Rhein induced apoptosis through the endoplasmic reticulum stress, caspase- and mitochondria-dependent pathways in SCC-4 human tongue squamous cancer cells. In Vivo 23: 309-316, 2009.
    OpenUrlAbstract/FREE Full Text
  28. ↵
    1. Lin SS,
    2. Huang HP,
    3. Yang JS,
    4. Wu JY,
    5. Hsia TC,
    6. Lin CC,
    7. Lin CW,
    8. Kuo CL,
    9. Gibson Wood W,
    10. Chung JG
    : DNA damage and endoplasmic reticulum stress mediated curcumin-induced cell cycle arrest and apoptosis in human lung carcinoma A-549 cells through the activation caspases cascade- and mitochondrial-dependent pathway. Cancer Lett 272: 77-90, 2008.
    OpenUrlCrossRefPubMed
  29. ↵
    1. Liotta LA,
    2. Stetler-Stevenson WG
    : Tumor invasion and metastasis: an imbalance of positive and negative regulation. Cancer Res 51: 5054s-5059s, 1991.
    OpenUrlAbstract/FREE Full Text
  30. ↵
    1. Zhang L,
    2. Shi J,
    3. Feng J,
    4. Klocker H,
    5. Lee C,
    6. Zhang J
    : Type IV collagenase (matrix metalloproteinase-2 and -9) in prostate cancer. Prostate Cancer Prostatic Dis 7: 327-332, 2004.
    OpenUrlPubMed
  31. ↵
    1. Guruvayoorappan C,
    2. Kuttan G
    : Amentoflavone inhibits experimental tumor metastasis through a regulatory mechanism involving MMP-2, MMP-9, prolyl hydroxylase, lysyl oxidase, VEGF, ERK-1, ERK-2, STAT-1, NM23 and cytokines in lung tissues of C57BL/6 mice. Immunopharmacol Immunotoxicol 30: 711-727, 2008.
    OpenUrlCrossRefPubMed
  32. ↵
    1. Monig SP,
    2. Baldus SE,
    3. Hennecken JK,
    4. Spiecker DB,
    5. Grass G,
    6. Schneider PM,
    7. Thiele J,
    8. Dienes HP,
    9. Holscher AH
    : Expression of MMP-2 is associated with progression and lymph node metastasis of gastric carcinoma. Histopathology 39: 597-602, 2001.
    OpenUrlCrossRefPubMed
  33. ↵
    1. Murray GI,
    2. Duncan ME,
    3. Arbuckle E,
    4. Melvin WT,
    5. Fothergill JE
    : Matrix metalloproteinases and their inhibitors in gastric cancer. Gut 43: 791-797, 1998.
    OpenUrlAbstract/FREE Full Text
  34. ↵
    1. Honda M,
    2. Mori M,
    3. Ueo H,
    4. Sugimachi K,
    5. Akiyoshi T
    : Matrix metalloproteinase-7 expression in gastric carcinoma. Gut 39: 444-448, 1996.
    OpenUrlAbstract/FREE Full Text
  35. ↵
    1. Yamashita K,
    2. Azumano I,
    3. Mai M,
    4. Okada Y
    : Expression and tissue localization of matrix metalloproteinase 7 (matrilysin) in human gastric carcinomas. Implications for vessel invasion and metastasis. Int J Cancer 79: 187-194, 1998.
    OpenUrlCrossRefPubMed
  36. ↵
    1. Kabashima A,
    2. Maehara Y,
    3. Kakeji Y,
    4. Baba H,
    5. Koga T,
    6. Sugimachi K
    : Clinicopathological features and overexpression of matrix metalloproteinases in intramucosal gastric carcinoma with lymph node metastasis. Clin Cancer Res 6: 3581-3584, 2000.
    OpenUrlAbstract/FREE Full Text
  37. ↵
    1. Nee L,
    2. Tuite N,
    3. Ryan MP,
    4. McMorrow T
    : TNF-alpha and IL-1 beta-mediated regulation of MMP-9 and TIMP-1 in human glome-rular mesangial cells. Nephron Exp Nephrol 107: e73-86, 2007.
    OpenUrlCrossRefPubMed
  38. ↵
    1. Zeisberg M,
    2. Maeshima Y,
    3. Mosterman B,
    4. Kalluri R
    : Renal fibrosis. Extracellular matrix microenvironment regulates migratory behavior of activated tubular epithelial cells. Am J Pathol 160: 2001-2008, 2002.
    OpenUrlCrossRefPubMed
  39. ↵
    1. Lee SO,
    2. Jeong YJ,
    3. Kim M,
    4. Kim CH,
    5. Lee IS
    : Suppression of PMA-induced tumor cell invasion by capillarisin via the inhibition of NF-kappaB-dependent MMP-9 expression. Biochem Biophys Res Commun 366: 1019-1024, 2008.
    OpenUrlCrossRefPubMed
  40. ↵
    1. Cho HJ,
    2. Kang JH,
    3. Kwak JY,
    4. Lee TS,
    5. Lee IS,
    6. Park NG,
    7. Nakajima H,
    8. Magae J,
    9. Chang YC
    : Ascofuranone suppresses PMA-mediated matrix metalloproteinase-9 gene activation through the Ras/Raf/MEK/ERK- and Ap1-dependent mechanisms. Carcinogenesis 28: 1104-1110, 2007.
    OpenUrlAbstract/FREE Full Text
    1. Zhang S,
    2. Li Z,
    3. Wu X,
    4. Huang Q,
    5. Shen HM,
    6. Ong CN
    : Methyl-3-indolylacetate inhibits cancer cell invasion by targeting the MEK1/2-ERK1/2 signaling pathway. Mol Cancer Ther 5: 3285-3293, 2006.
    OpenUrlAbstract/FREE Full Text
  42. ↵
    1. Chakraborti S,
    2. Mandal M,
    3. Das S,
    4. Mandal A,
    5. Chakraborti T
    : Regulation of matrix metalloproteinases: an overview. Mol Cell Biochem 253: 269-285, 2003.
    OpenUrlCrossRefPubMed
  43. ↵
    1. Simon C,
    2. Simon M,
    3. Vucelic G,
    4. Hicks MJ,
    5. Plinkert PK,
    6. Koitschev A,
    7. Zenner HP
    : The p38 SAPK pathway regulates the expression of the MMP-9 collagenase via AP-1-dependent promoter activation. Exp Cell Res 271: 344-355, 2001.
    OpenUrlCrossRefPubMed
  44. ↵
    1. Woo JH,
    2. Lim JH,
    3. Kim YH,
    4. Suh SI,
    5. Min DS,
    6. Chang JS,
    7. Lee YH,
    8. Park JW,
    9. Kwon TK
    : Resveratrol inhibits phorbol myristate acetate-induced matrix metalloproteinase-9 expression by inhibiting JNK and PKC delta signal transduction. Oncogene 23: 1845-1853, 2004.
    OpenUrlCrossRefPubMed
  45. ↵
    1. Kwaan HC
    : The plasminogen-plasmin system in malignancy. Cancer Metastasis Rev 11: 291-311, 1992.
    OpenUrlCrossRefPubMed
  46. ↵
    1. Duffy MJ,
    2. Maguire TM,
    3. McDermott EW,
    4. O'Higgins N
    : Urokinase plasminogen activator: a prognostic marker in multiple types of cancer. J Surg Oncol 71: 130-135, 1999.
    OpenUrlCrossRefPubMed
  47. ↵
    1. Chen PN,
    2. Hsieh YS,
    3. Chiou HL,
    4. Chu SC
    : Silibinin inhibits cell invasion through inactivation of both PI3K-Akt and MAPK signaling pathways. Chem Biol Interact 156: 141-150, 2005.
    OpenUrlCrossRefPubMed
    1. Lee SJ,
    2. Park SS,
    3. Lee US,
    4. Kim WJ,
    5. Moon SK
    : Signaling pathway for TNF-alpha-induced MMP-9 expression: mediation through p38 MAP kinase, and inhibition by anticancer molecule magnolol in human urinary bladder cancer 5637 cells. Int Immunopharmacol 8: 1821-1826, 2008.
    OpenUrlCrossRefPubMed
  49. ↵
    1. Sun LC,
    2. Luo J,
    3. Mackey LV,
    4. Fuselier JA,
    5. Coy DH
    : A conjugate of camptothecin and a somatostatin analog against prostate cancer cell invasion via a possible signaling pathway involving PI3K/Akt, alphaVbeta3/alphaVbeta5 and MMP-2/-9. Cancer Lett 246: 157-166, 2007.
    OpenUrlCrossRefPubMed
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Phenethyl Isothiocyanate Inhibits Migration and Invasion of Human Gastric Cancer AGS Cells through Suppressing MAPK and NF-κB Signal Pathways
MEI-DUE YANG, KUANG-CHI LAI, TUNG-YUAN LAI, SHU-CHUN HSU, CHAO-LIN KUO, CHUN-SHU YU, MENG-LIANG LIN, JAI-SING YANG, HSIU-MAAN KUO, SHIN-HWAR WU, JING-GUNG CHUNG
Anticancer Research Jun 2010, 30 (6) 2135-2143;
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