Fucoxanthin derivatives from Sarggassum siliquastrum inhibit matrix metalloproteinases by suppressing NF-κB and MAPKs in human fibrosarcoma cells

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  • ABSTRACT

    Fucoxanthin is known to be an effective cell proliferation inhibitor with anti-tumor and anti-angiogenic activities. However, there is a lack of data regarding the biological effects of cis isomers of fucoxanthin. To assess the potential therapeutic properties of 9′-cis-(6′R) fucoxanthin (FcA), and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) isolated from Sarggassum siliquastrum, we investigated their inhibitory effects on matrix metalloproteinases (MMPs) in phorbol 12-myristate 13-acetate (PMA)-induced human fibrosarcoma (HT1080) cells. FcA and FcB reduced MMP-2 and MMP-9 protein and mRNA levels, as well as the migration of these cells, in a dose-dependent manner. Additionally, FcA and FcB increased levels of MMPs inhibition factors such as tissue inhibitor of metalloproteinase-1. FcA and FcB significantly inhibited the transcriptional activity of nuclear factor кB (NF-кB) and by inhibiting c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinases. Our results demonstrate that suppression of the NF-κB, JNK, and p38 signaling pathways may inhibit PMA-induced MMP-2 and MMP-9 activity. Therefore, FcA and FcB may be useful in noninvasive therapeutic strategies against fibrosarcoma metastasis.


  • KEYWORD

    fucoxanthin , human fibrosarcoma cells (HT1080) , matrix metalloproteinases (MMPs) , mitogen-activated protein kinase (MAPK) , nuclear factor-kappaB (NF-κB) , Sargassum siliquastrum

  • 1. Bauvois B. 2012 New facets of matrix metalloproteinases MMP-2 and MMP-9 as cell surface transducers: outside-in signaling and relationship to tumor progression [Biochim. Biophys. Acta] Vol.1825 P.29-36 google
  • 2. Chou Y. C., Sheu J. R., Chung C. L., Chen C. Y., Lin F. L., Hsu M. J., Kuo Y. H., Hsiao G. 2010 Nuclear-targeted inhibition of NF-kB on MMP-9 production by N-2-(4-bromophenyl) ethyl caffeamide in human monocytuc cells [Chem. Biol. Interact.] Vol.184 P.403-412 google doi
  • 3. Debret R., Brassart-Pasco S., Lorin J., Martoriati A., Deshorgue A., Maquart F.-X., Hornebeck W., Rahman I., Antonicelli F. 2008 Ceramide inhibition of MMP-2 expression and human cancer bronchial cell invasiveness involve decreased histone acetylation [Biochim. Biophys. Acta] Vol.1783 P.1718-1727 google doi
  • 4. Fung A., Hamid N., Lu J. 2013 Fucoxanthin content and antioxidant properties of Undaria pinnatifida [Food Chem] Vol.136 P.1055-1062 google doi
  • 5. Halliwell B., Gutteridge J. M. C., Halliwell B., Gutteridge J. M. C. 1999 Antioxidant defenses P.105-109 google
  • 6. Hansen M. B., Nielsen S. E., Berg K. 1989 Re-examination and further development of a precise and rapid dye method for measuring cell growth/cell kill [J. Immunol. Methods] Vol.119 P.203-210 google doi
  • 7. Hanigan C. L., van Engeland M., De Bruine A. P., Wouters K. A., Weijenberg M. P., Eshleman J. R., Herman J. G. 2008 An inactivating mutation in HDAC2 leads to dysregulation of apoptosis mediated by APAF1 [Gastroenterology] Vol.135 P.1654-1664 google doi
  • 8. Heo S.-J., Jeon Y.-J. 2009 Protective effect of fucoxanthin isolated from Sargassum siliquastrumon on UV-B induced cell damage [J. Photochem. Photobiol. B] Vol.95 P.101-107 google doi
  • 9. Heo S.-J., Ko S.-C., Kang S.-M., Kang H.-S., Kim J.-P., Kim S.-H., Lee K.-W., Cho M.-G., Jeon Y.-J. 2008 Cytoprotective effect of fucoxanthin isolated from brown algae Sargassum siliquastrum against H2O2-induced cell damage [Eur. Food Res. Technol.] Vol.228 P.145-151 google doi
  • 10. Heo S.-J., Yoon W.-J., Kim K.-N., Ahn G.-N., Kang S.-M., Kang D.-H., Affan A., Oha C., Jung W.-K., Jeon Y.-J. 2010 Evaluation of anti-inflammatory effect of fucoxanthin isolated from brown algae in lipopolysaccharidestimulated RAW 264.7 macrophages [Food Chem. Toxicol.] Vol.48 P.2045-2051 google doi
  • 11. Heo S.-J., Yoon W.-J., Kim K.-N., Oh C., Choi Y.-U., Yoon K.-T., Kang D.-H., Qian Z.-J., Choi I.-W., Jung W.-K. 2012 Anti-inflammatory effect of fucoxanthin derivatives isolated from Sargassum siliquastrum in lipopolysaccharide-stimulated RAW 264.7 macrophage [Food Chem. Toxicol.] Vol.50 P.3336-3342 google doi
  • 12. Hosokawa M., Kudo M., Maeda H., Kohno H., Tanaka T., Miyashita K. 2004 Fucoxanthin induces apoptosis and enhances the anti-proliferative effect of the PPARγ ligand troglitazone, on colon cancer cells [Biochim. Biophys. Acta] Vol.1675 P.113-119 google doi
  • 13. Hwang Y. P., Yun H. J., Kim H. G., Han E. H., Lee G. W., Jeong H. G. 2010 Suppression of PMA-induced tumor cell invasion by dihydroartemisinin via inhibition of PKCα/Raf/MAPKs and NF-kB/AP-1-dependent mechanisms [Biochem. Pharmacol.] Vol.79 P.1714-1726 google doi
  • 14. Jang K. H., Lee B. H., Choi B. W., Lee H.-S., Shin J. 2005 Chromenes from the brown alga Sargassum siliquastrum [J. Nat. Prod.] Vol.68 P.716-723 google doi
  • 15. Kang M.-C., Kim E.-A., Kang S.-M., Wijesinghe W. A. J. P., Yang X., Kang N., Jeon Y.-J. 2012 Thermostability of a marine polyphenolic antioxidant dieckol, derived from the brown seaweed Ecklonia cava [Algae] Vol.27 P.205-213 google doi
  • 16. Kang S.-I., Ko H.-C., Shin H.-S., Kim H.-M., Hong Y.-S., Lee N.-H., Kim S.-J. 2011 Fucoxanthin exerts differing effects on 3T3-L1 cells according to differentiation stage and inhibits glucose uptake in mature adipocytes [Biochem. Biophys. Res. Commun.] Vol.409 P.769-774 google doi
  • 17. Kaomongkolgit R., Cheepsunthorn P., Pavasant P., Sanchavanakit N. 2008 Iron increases MMP-9 expression through activation of AP-1 via ERK/Akt pathway in human head and neck squamous carcinoma cells [Oral Oncol.] Vol.44 P.587-594 google doi
  • 18. Kim B. S., Park J.-Y., Kang H.-J., Kim H.-J., Lee J. 2014 Fucoidan/FGF-2 induces angiogenesis through JNKand p38-mediated activation of AKT/MMP-2 signalling [Biochem. Biophys. Res. Commun.] Vol.450 P.1333-1338 google doi
  • 19. Kim K.-N., Ahn G., Heo S.-J., Kang S.-M., Kang M.-C., Yang H.-M., Kim D., Roh S. W., Kim S.-K., Jeon B.-T., Park P.-J., Jung W.-K., Jeon Y. J. 2013 Inhibition of tumor growth in vitro and in vivo by fucoxanthin against melanoma B16F10 cells [Environ. Toxicol. Pharmacol.] Vol.35 P.39-46 google doi
  • 20. Kim K.-N., Heo S.-J., Yoon W.-J., Kang S.-M., Ahn G., Yi T.-H., Jeon Y.-J. 2010 Fucoxanthin inhibits the inflammatory response by suppressing the activation of NF-κB and MAPKs in lipopolysaccharide-induced RAW 264.7 macrophages [Eur. J. Pharmacol.] Vol.649 P.369-375 google doi
  • 21. Kim M.-M., Ta Q. V., Mendis E., Rajapakse N., Jung W.-K., Byun H.-G., Jeon Y.-J., Kim S.-K. 2006 Phlorotannins in Ecklonia cava extract inhibit matrix metalloproteinase activity [Life Sci] Vol.79 P.1436-1443 google doi
  • 22. Kong C.-S., Kim J.-A., Ahn B., Byun H.-G., Kim S.-K. 2010 Carboxymethylations of chitosan and chitin inhibit MMP expression and ROS scavenging in human fibrosarcoma cells [Process Biochem] Vol.45 P.179-186 google doi
  • 23. Kong C.-S., Kim Y. A., Kim M.-M., Park J.-S., Kim J.-A., Kim S.-K., Lee B.-J., Nam T. J., Seo Y. 2008 Flavonoid glycosides isolated from Salicornia herbacea inhibit matrix metalloproteinase in HT1080 cells [Toxicol. In Vitro] Vol.22 P.1742-1748 google doi
  • 24. Kwak H.-J., Park M.-J., Cho H., Park C.-M., Moon S.-I., Lee H.-C., Park I.-C., Kim M.-S., Rhee C.-H., Hong S.-I. 2006 Transforming growth factor-β1 induces tissue inhibitor of metalloproteinase-1 expression via activation of extracellular signal-regulated kinase and Sp1 in human fibrosarcoma cells [Mol. Cancer Res.] Vol.4 P.209-220 google doi
  • 25. Lee S.-K., Yang J. Y., Han D. C., Son K.-H., Kwon B.-M., Chun H. K. 2007 Inhibitory effect of obovatal on the migration and invasion of HT1080 cells via the inhibition of MMP-2 [Bioorg. Med. Chem.] Vol.15 P.4085-4090 google doi
  • 26. Maeda H., Tsukui T., Sashima T., Hosokawa M., Miyashita K. 2008 Seaweed carotenoid, fucoxanthin, as a multifunctional nutrient [Asia Pac. J. Clin. Nutr.] Vol.17 P.196-199 google
  • 27. McCawley L. J., Matrisian L. M. 1999 Matrix metalloproteinases: multifunctional contributors to tumor progression [Mol. Med. Today] Vol.6 P.149-156 google
  • 28. Mendis E., Kim M.-M., Rajapakse N., Kim S.-K. 2006 Carboxy derivatized glucosamine is a potent inhibitor of matrix metalloproteinase-9 in HT1080 cells [Bioorg. Med. Chem. Lett.] Vol.16 P.3105-3110 google doi
  • 29. Mendis E., Kim M.-M., Rajapakse N., Kim S.-K. 2009 The inhibitory mechanism of a novel cationic glucosamine derivative against MMP-2 and MMP-9 expressions [Bioorg. Med. Chem. Lett.] Vol.19 P.2755-2759 google doi
  • 30. Miyashita K., Nishikawa S., Beppu F., Tsukui T., Abea M., Hosokawa M. 2011 The allenic carotenoid fucoxanthin, a novel marine nutraceutical from brown seaweeds [J. Sci. Food Agric.] Vol.91 P.1166-1174 google doi
  • 31. Nakazawa Y., Sashima T., Hosokawa M., Miyashita K. 2009 Comparative evaluation of growth inhibitory effect of stereoisomers of fucoxanthin in human cancer cell lines [J. Funct. Food] Vol.1 P.88-97 google doi
  • 32. Nguyen V.-T., Qian Z.-J., Ryu B. M., Kim K.-N., Kim D., Kim Y.-M., Jeon Y.-J., Park W. S., Choi I.-W., Kim G. H., Je J.-Y., Jung W.-K. 2013 Matrix metalloproteinases (MMPs) inhibitory effects of an octameric oligopeptide isolated from abalone Haliotis discus hannai [Food Chem] Vol.141 P.503-509 google doi
  • 33. Park H. J., Chung H.-J., Min H.-Y., Park E.-J., Hong J.-Y., Kim W. B., Kim S. H., Lee S. K. 2005 Inhibitory effect of DA-125, a new anthracyclin analog antitumor agent, on the invasion of human fibrosarcoma cells by down-regulating the matrix metalloproteinases [Biochem. Pharmacol.] Vol.71 P.21-31 google doi
  • 34. Park I.-H., Kim M.-M. 2012 Spermidine inhibits MMP-2 via modulation of histone acetyltransferase and histone deacetylase in HDFs [Int. J. Biol. Macromol.] Vol.51 P.1003-1007 google doi
  • 35. Rajapakse N., Kim M. M., Mendis M., Huang R., Kim S. K. 2006 Carboxylated chitooligosaccharides (CCOS) inhibit MMP-9 expression in human fibrosarcoma cells via down-regulation of AP-1 [Biochim. Biophys. Acta] Vol.1760 P.1780-1788 google doi
  • 36. Shiratori K., Ohgami K., Ilieva I., Jin X.-H., Koyama Y., Miyashita K., Yoshida K., Kase S., Ohno S. 2005 Effects of fucoxanthin on lipopolysaccharide-induced inflammation in vitro and in vivo [Exp. Eye Res.] Vol.81 P.422-428 google doi
  • 37. Sivathanu B., Palaniswamy S. 2012 Purification and characterization of carotenoids from green algae Chlorococcum humicola by HPLC-NMR and LC-MS-APCI [Biomed. Prev. Nutr.] Vol.2 P.276-282 google doi
  • 38. Yan X., Chuda Y., Suzuki M., Nagata T. 1999 Fucoxanthin as the major antioxidant in Hijikia fusiformis, a common edible seaweed [Biosci. Biotechnol. Biochem.] Vol.63 P.605-607 google doi
  • 39. Yu R.-X., Hu X.-M., Xu S.-Q., Jiang Z.-J., Yang W. 2011 Effects of fucoxanthin on proliferation and apoptosis in human gastric adenocarcinoma MGC-803 cells via JAK/STAT signal pathway [Eur. J. Pharmacol.] Vol.657 P.10-9 google doi
  • 40. Zhao Z., Huang G., Wang B., Zhong Y. 2013 Inhibition of NF-kappaB activation by pyrrolidine dithiocarbamate partially attenuates hippocampal MMP-9 activation and improves cognitive deficits in streptozotocin-induced diabetic rats [Behav. Brain Res.] Vol.23 P.44-47 google
  • [Fig. 1.] Chemical structures of fucoxnathin 9′-cis-(6′R)-isomer (FcA) (A), and 13-cis-(6′R) isomer (B), and 13′-cis-(6′R) isomer complex (FcB) (C) isolated from Sargassum siliquastrum.
    Chemical structures of fucoxnathin 9′-cis-(6′R)-isomer (FcA) (A), and 13-cis-(6′R) isomer (B), and 13′-cis-(6′R) isomer complex (FcB) (C) isolated from Sargassum siliquastrum.
  • [Fig. 2.] Cytotoxic effects of fucoxanthin derivatives (9′-cis-(6′R) fucoxanthin [FcA] and 13-cis and 13′-cis-(6′R) fucoxanthin complex [FcB]) on HT1080 cells in the presence and absence of fetal bovine serum (FBS). Cells were treated for 24 h with various concentrations (5, 10, and 50 μM) of FcA (A) and FcB (B). Cytotoxicity was determined using the MTT assay. Values are expressed as mean ± standard deviation of triplicate experiments.
    Cytotoxic effects of fucoxanthin derivatives (9′-cis-(6′R) fucoxanthin [FcA] and 13-cis and 13′-cis-(6′R) fucoxanthin complex [FcB]) on HT1080 cells in the presence and absence of fetal bovine serum (FBS). Cells were treated for 24 h with various concentrations (5, 10, and 50 μM) of FcA (A) and FcB (B). Cytotoxicity was determined using the MTT assay. Values are expressed as mean ± standard deviation of triplicate experiments.
  • [Fig. 3.] Inhibition of HT1080 cells migration by 9′-cis-(6′R) fucoxanthin (FcA) (A) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) (B). Cells were grown in 6-well plate and confluent monolayer were wounded and then FcA and FcB (5, 10, and 50 μM) were added and incubated for 24 h. After injury line was made on the confluent monolayer of cells. At 0 and 24 h, the cells were photographed under a phase contrast microscope.
    Inhibition of HT1080 cells migration by 9′-cis-(6′R) fucoxanthin (FcA) (A) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) (B). Cells were grown in 6-well plate and confluent monolayer were wounded and then FcA and FcB (5, 10, and 50 μM) were added and incubated for 24 h. After injury line was made on the confluent monolayer of cells. At 0 and 24 h, the cells were photographed under a phase contrast microscope.
  • [Fig. 4.] Effect of 9′-cis-(6′R) fucoxanthin (FcA) (A) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) (B) on the gelatinolytic activity of matrix metalloproteinase (MMP)-2 and MMP-9 in HT1080 cell line determined by gelatin zymography. Gelatinolytic activities of MMP-2 and MMP-9 in conditioned serum-free media were detected by electrophoresis on gelatin containing 10% polyacrylamide gel. Areas and relative intensities of gelatin digested bands were quantified by densitometry and expressed as relative MMP-2 and MMP-9 activities compared to those of phorbol 12-myristate 13-acetate (PMA) alone treated cells. Values are expressed as mean ± standard deviation of triplicate experiments. *p < 0.05 indicates significant differences from the PMA-stimulated group.
    Effect of 9′-cis-(6′R) fucoxanthin (FcA) (A) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) (B) on the gelatinolytic activity of matrix metalloproteinase (MMP)-2 and MMP-9 in HT1080 cell line determined by gelatin zymography. Gelatinolytic activities of MMP-2 and MMP-9 in conditioned serum-free media were detected by electrophoresis on gelatin containing 10% polyacrylamide gel. Areas and relative intensities of gelatin digested bands were quantified by densitometry and expressed as relative MMP-2 and MMP-9 activities compared to those of phorbol 12-myristate 13-acetate (PMA) alone treated cells. Values are expressed as mean ± standard deviation of triplicate experiments. *p < 0.05 indicates significant differences from the PMA-stimulated group.
  • [Fig. 5.] Inhibitory effect of phorbol 12-myristate 13-acetate (PMA)-stimulated matrix metalloproteinase (MMP)-2, MMP-9 and tissue inhibitor of metalloproteinase 1 (TIMP-1) mRNA (A) and protein (B) expression by 9′-cis-(6′R) fucoxanthin (FcA) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) in HT10880 cells. (A) HT1080 cells were pre-incubated for 18 h, and the cells were stimulated with PMA (10 ng mL?1) for 24 h in the presence of FcA and FcB (5, 10, and 50 μM). Cell lysates were electrophoresed, and the expression levels of MMP-2, MMP-9, and TIMP-1 were detected with specific antibodies. (B) After PMA treatment, total RNA was prepared from HT10880 cells and reverse transcriptase-polymerase chain reaction was preformed for the MMP-2, MMP-9, and TIMP-1 genes. β-Actin and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were used as internal control for Western blot analysis and reverse transcriptase-polymerase chain reaction assays. Values are expressed as mean ± standard deviation of triplicate experiments. *p < 0.05 and **p < 0.01 indicate significant differences from the PMA-stimulated group.
    Inhibitory effect of phorbol 12-myristate 13-acetate (PMA)-stimulated matrix metalloproteinase (MMP)-2, MMP-9 and tissue inhibitor of metalloproteinase 1 (TIMP-1) mRNA (A) and protein (B) expression by 9′-cis-(6′R) fucoxanthin (FcA) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) in HT10880 cells. (A) HT1080 cells were pre-incubated for 18 h, and the cells were stimulated with PMA (10 ng mL?1) for 24 h in the presence of FcA and FcB (5, 10, and 50 μM). Cell lysates were electrophoresed, and the expression levels of MMP-2, MMP-9, and TIMP-1 were detected with specific antibodies. (B) After PMA treatment, total RNA was prepared from HT10880 cells and reverse transcriptase-polymerase chain reaction was preformed for the MMP-2, MMP-9, and TIMP-1 genes. β-Actin and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were used as internal control for Western blot analysis and reverse transcriptase-polymerase chain reaction assays. Values are expressed as mean ± standard deviation of triplicate experiments. *p < 0.05 and **p < 0.01 indicate significant differences from the PMA-stimulated group.
  • [Fig. 6.] Inhibitory effect of 9′-cis-(6′R) fucoxanthin (FcA) (A) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) (B) on the protein level of p38 and c-Jun N-terminal kinase (JNK) in HT1080 cells. HT1080 cells were pre-incubated for 18 h, and the macrophages were stimulated with phorbol 12-myristate 13-acetate (PMA; 10 ng mL?1) in the presence of FcA and FcB (5, 10, and 50 μM). The levels of p-JNK, JNK, p-p38, and p38 were determined via Western blotting. Values are expressed as mean ± standard deviation of triplicate experiments. *p < 0.05 and **p < 0.01 indicate significant differences from the PMA-stimulated group.
    Inhibitory effect of 9′-cis-(6′R) fucoxanthin (FcA) (A) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) (B) on the protein level of p38 and c-Jun N-terminal kinase (JNK) in HT1080 cells. HT1080 cells were pre-incubated for 18 h, and the macrophages were stimulated with phorbol 12-myristate 13-acetate (PMA; 10 ng mL?1) in the presence of FcA and FcB (5, 10, and 50 μM). The levels of p-JNK, JNK, p-p38, and p38 were determined via Western blotting. Values are expressed as mean ± standard deviation of triplicate experiments. *p < 0.05 and **p < 0.01 indicate significant differences from the PMA-stimulated group.
  • [Fig. 7.] Inhibitory effect of 9′-cis-(6′R) fucoxanthin (FcA) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) on phorbol 12-myristate 13-acetate (PMA)-stimulated activation of nuclear factor κB (NF-κB) in HT1080 cells. HT1080 cells were pre-incubated for 18 h, and the cells were stimulated with PMA (10 ng mL?1) in the presence of FcA and FcB (5, 10, and 50 μM). (A) The cytosolic and nuclear extracts were prepared as described in section 2 and evaluated for NF-κB p65 via Western blot analysis. Values are expressed as mean ± standard deviation of triplicate experiments. *p < 0.05 and **p < 0.01 indicate significant differences from the PMA-stimulated group. (B) The NF-κB p65 expression was observed by immunofluorescence staining in HT1080 cells. Stained nuclei with DAPI solution were then photographed with a fluorescent microscope using a blue filter.
    Inhibitory effect of 9′-cis-(6′R) fucoxanthin (FcA) and 13-cis and 13′-cis-(6′R) fucoxanthin complex (FcB) on phorbol 12-myristate 13-acetate (PMA)-stimulated activation of nuclear factor κB (NF-κB) in HT1080 cells. HT1080 cells were pre-incubated for 18 h, and the cells were stimulated with PMA (10 ng mL?1) in the presence of FcA and FcB (5, 10, and 50 μM). (A) The cytosolic and nuclear extracts were prepared as described in section 2 and evaluated for NF-κB p65 via Western blot analysis. Values are expressed as mean ± standard deviation of triplicate experiments. *p < 0.05 and **p < 0.01 indicate significant differences from the PMA-stimulated group. (B) The NF-κB p65 expression was observed by immunofluorescence staining in HT1080 cells. Stained nuclei with DAPI solution were then photographed with a fluorescent microscope using a blue filter.