• Radiation Oncology Journal
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• v.33 no.4
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• pp.328-336
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• 2015

Purpose: Past studies have reported that S-allylcysteine (SAC) inhibits the migration and invasion of cancer cells through the restoration of E-cadherin, the reduction of matrix metalloproteinase (MMP) and Slug protein expression, and inhibition of the production of reactive oxygen species (ROS). Furthermore, evidence is emerging that shows that ROS induced by radiation could increase Met activation. Following on these reports of SAC and Met, we investigated whether SAC could suppress Met activation. Materials and Methods: Wound healing, invasion, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium (MTT), soft agar colony forming, western blotting, and gelatin zymography assays were performed in the human nasopharyngeal cancer cell lines HNE1 and HONE1 treated with SAC (0, 10, 20, or 40 mM) and hepatocyte growth factor (HGF). Results: This study showed that SAC could suppress the migration and invasion of HNE1 and HONE1 cell lines by inhibiting p-Met. An increase of migration and invasion induced by HGF and its decrease in a dose dependent manner by SAC in wound healing and invasion assays was observed. The reduction of p-Met by SAC was positively correlated with p-focal adhesion kinase (p-FAK) and p-extracellular related kinase (p-ERK in both cell lines). SAC reduced Slug, MMP2, and MMP9 involved in migration and invasion with the inhibition of Met-FAK signaling. Conclusion: These results suggest that SAC inhibited not only Met activation but also the downstream FAK, Slug, and MMP expression. Finally, SAC may be a potent anticancer compound for nasopharyngeal cancer treated with radiotherapy.

• Journal of Embryo Transfer
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• v.33 no.3
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• pp.99-105
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• 2018

To determine the differences in the in-vitro ovum maturation process of bovine, we compared the expression of MMPs in these oocytes and cumulus cell throughout oocytes maturated. In an attempt to investigate the effect of MMP activation and inhibitors in total protein of cumulus cell and, oocytes during oocytes maturation, we examined and monitored the localization and expression of MMPs (MMP-2 and MMP-9), TIMPs (TIMP-2 and TIMP-3), as well as their expression profiles (Real-time PCR, Gelatin Zymography and ELISA). Our results that the bovine oocytes MMP-2 and MMP-9 level was significantly associated with the rate of maturity of oocytes (P<0.05). In cumulus cell, MMP-2 was highly expressed in all stages of the oocyte's maturation. The final oocytes maturation exhibited strong gelatinase activity. There was no significant correlation between cumulus cell MMP-9 and the maturation rate of oocytes. However, for the oocyte cytoplasm MMP-9 expression was significant correlation to the maturation oocytes. There was no significant correlation between cumulonimbus cells MMP-9 and oocyte maturation rates; however, for oocyte cytoplasm, MMP-9 expression was significantly correlated with mature oocyte. However, the TIMP-1 and TIMP-2 protein expression patterns are not correlated with the maturation rate of the oocyte. Our results suggest that MMP different expression pattern may regulate the morphological remodeling of oocyte's in the cumulus cell. Further, the MMP-2 expression has a strong relation with a higher maturation rate of the oocyte.

• Journal of Physiology & Pathology in Korean Medicine
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• v.18 no.6
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• pp.1602-1607
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• 2004

3-O-D-galactopyranosylglyceride (GPG; fatty acids R1, R2 = myristic acid 11.62%, palmitic acid 61.90% and oleic acid 26.48%) was isolated from Chrysanthemum coronarium L that has been used for treating renal and cardiovascular diseases as one of vegetables or medicinal drug. However, little was known about the anti-angiogenic activity of GPG. Thus, anti-angiogenic effect of GPG was evaluated in human umbilical vein endothelial cells (HUVECs) in vitro and in vivo. GPG effectively inhibited bFGF-induced migration and invasion of HUVECs in a concentration-dependent manner, whereas it did not inhibit bFGF-induced proliferation and capillary-like tube formation of HUVECs. To examine the mechanism of anti-angiogenic activity of GPG, gelatin zymography was carried out. GPG downregulated the expression of matrix metalloproteinase-2 in a concentration-dependent manner. Furthermore, GPG significantly disrupted bFGF-induced neovascularization on the chick chorioallantoic membrane assay in vivo. These results suggest that 3-O--D-galactopyranosylglyceride may inhibit neovascularization by inhibiting angiogenic activity of endothelial cells via regulation of matrix metalloproteinase-2 (MMP-2).

• Parasites, Hosts and Diseases
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• v.48 no.2
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• pp.151-155
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• 2010

Allergen extracts from dust mites and cockroaches commonly found in Korean homes were used to evaluate their enzymatic activity as they are believed to influence allergenicity. Allergen extracts were prepared from 3 dust mite species (Dermatophagoides farinae, D. pteronyssinus, and Tyrophagus putrescentiae) and 3 cockroach species (Blattella germanica, Periplaneta americana, and P. fuliginosa) maintained in the Korea National Arthropods of Medical Importance Resource Bank. Proteins were extracted in PBS after homogenization using liquid nitrogen. The activities of various enzymes were investigated using the API Zym system. No significant difference in phosphatase, lipase, or glycosidase activity was observed among the 6 allergen extracts, but much difference was observed in protease activity. Protease activity was assessed in more detail by gelatin zymography and the EnzChek assay. Extract from T. putrescentiae showed the highest protease activity, followed by those of the cockroach extracts. Extracts from D. farinae and D. pteronyssinus showed only weak protease activity. Gelatinolytic activity was detected mainly in a 30-kDa protein in D. farinae, a 28-kDa protein in D. pteronyssinus, a > 26-kDa protein in T. putrescentiae, a > 20-kDa protein in B. germanica, and a > 23-kDa protein in P. americana and P. fuliginosa. The information on various enzymatic activities obtained in this study may be useful for future studies. In particular, the strong protease activity found in cockroach extracts could contribute to sensitization to cockroach allergens, which is known to be associated with the development of asthma.

• Journal of Microbiology and Biotechnology
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• v.16 no.12
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• pp.1904-1911
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• 2006

The cell-free extracts of 250 yeasts were screened for their in vitro anti-angiogenic activity, to develop a new cancer metastasis inhibitor. Saccharomyces cerevisiae K-7 was selected as the producer of the anti-angiogenic agent, because it had the highest anti-angiogenic activity. The anti-angiogenic agent was produced maximally from hydrolysates of Saccharomyces cerevisiae K-7, when the yeast was cultured in yeast extract-peptone-dextrose medium at 30$^{\circ}C$ for 24 h, and cell-free extracts were than digested with pepsin for 4 h at 37$^{\circ}C$. The anti-angiogenic agent was further purified by ultrafiltration, Sephadex G-25 gel permeation chromatography and reverse-phase HPLC, and the anti-angiogenic activity of the final purified preparation was 72.7% at 10 $\mu$M/egg. The purified anti-angiogenic agent was found to originate from the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) molecule of Saccharomyces cerevisiae K-7, and its peptide sequence was Val-Ser-Trp-Tyr-Asp-Asn-Glu-Tyr-Gly-Tyr-Ser-Thr-Arg-Val-Val-Asp. In the MTT assay, the shape of the HT-l 080 cell was clearly changed to a circular type at 0.2 mM purified anti-angiogenic agent. This result indicated that the growth of the HT-I080 cell was significantly inhibited at 0.2 mM of the purified anti-angiogenic agent. The MMP activity of the treated HT-l080 cells was not affected, evidenced by the gelatin zymography, indicating that the anti-angiogenic mechanism of the purified anti-angiogenic agent is not mediated through MMP activity.

• Journal of Microbiology and Biotechnology
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• v.23 no.11
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• pp.1536-1543
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• 2013

Proteases produced by Xenorhabdus are known to play a significant role in virulence leading to insect mortality. The present study was undertaken to purify and characterize protease from Xenorhabdus indica, an endosymbiont of nematode Steinernema thermophilum, and to decipher its role in insect mortality and its efficacy to control Helicoverpa armigera. A set of 10 strains of Xenorhabdus isolated from different regions of India were screened for protease activity on the basis of zone of clearing on gelatin agar plates. One potent strain of Xenorhabdus indica was selected for the production of protease, and the highest production (1,552 U/ml) was observed at 15-18 h of incubation at $28^{\circ}C$ in soya casein digest broth. The extracellular protease was purified from culture supernatant using ammonium sulfate precipitation and ion-exchange chromatography. The enzyme was further characterized by SDS-PAGE and zymography, which confirmed the purity of the protein and its molecular mass was found to be ~52 kDa. Further MALDI-TOF/TOF analysis and effect of metal chelating agent 1,10-phenanthrolin study revealed the nature of the purified protease as a secreted alkaline metalloprotease. The bioefficacy of the purified protease was also tested against cotton bollworm (Helicoverpa armigera) and resulted in $67.9{\pm}0.64%$ mortality within one week. This purified protease has the potential to be developed as a natural insecticidal agent against a broad range of agriculturally important insects.

• Nutritional Sciences
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• v.7 no.3
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• pp.144-150
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• 2004

We have investigated the effect of calcium spirulan(Ca-SP) isolated from a blue-green alga Spirulina platensis, which is a sulfated polysaccharide chelating calcium and mainly composed of rhamnose and fructose, on invasion of both B16- BL6 melanoma cells, Colon 26 carcinoma and HT-1080 fibrosarcoma cells through reconstituted basement membrane (Matrigel). Ca-SP significantly inhibited the invasion of these tumor cells through Matrigel/fibronectin-coated filters in a concentration-dependent manner. Ca-SP also inhibited the haptotactic migration of tumor cells to laminin, but it had no inhibitory effect on tumor cell migration to fibronectin-coated filters. Ca-SP prevented the adhesion of B16-BL6 cells to Matrigel- and laminin-substrates but did not affect the adhesion to fibronectin. The pretreatment of tumor cells with Ca-SP inhibited the adhesion to laminin in a concentration-dependent fashion, while the pretreatment of laminin-substrates did not. Ca-SP had no effect on the production and activation of type IV collagenase in gelatin zymography. In contraset, Ca-SP significantly inhibited degradation of heparan sulfate by purified heparanase. The experimental lung metastasis was significantly reduced by co-injection of B16-BL6 cells with Ca-SP in a dose-dependent manner. Seven intermittent ⅰ.ⅴ. injection of 100$\mu\textrm{g}$ of Ca-SP caused a marked decrease of lung tumor colonization of B16-BL6 cells in a spontaneous lung metastasis model. These results suggest that Ca-SP, a novel sulfated polysaccharide, could reduce the lung colonization of B16-BL6 melanoma cells in experimental metastasis model, by inhibiting the tumor invasion of basement membrane Matrigel, probably through the prevention of the adhesion and migration of tumor cells to laminin-substrate and of the heparanase activity.

• Biomolecules & Therapeutics
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• v.12 no.1
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• pp.1-8
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• 2004

In chronic airway inflammatory diseases such as asthma and chronic bronchitis, it has been suggested that matrix metalloproteinases secreted from infiltrating neutrophil contribute the pathogenesis of the disease and have been a focus of intense investigation. We report here that hamster tracheal surface epithelial goblet cells (HTSE cells) produce matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of metalloproteinase-2 (TIMP-2). Matrix metalloproteinase activities were investigated using [$^3H$]collagen-digestion assay and gelatin zymography. The subtype of matrix metalloproteinases expressed from HTSE cells was MMP-2 (gelatinase A), which was determined by Western blot with various subtype selective anti-matrix metalloproteinase antibodies. The MMP-2 and TIMP-2 cDNAs from HTSE cells were partially cloned by RT-PCR and they reveal more than 90% of sequence homology with those from human, rat and mouse. The collagenolytic activity was increased with the secretory differentiation of the HTSE cell and it was found that zymogen activation was responsible for the increased MMP-2 activity in HTSE cells. The results from the present study suggest that the metaplastic secretory differentiation of airway goblet cells may affect chronic airway inflammatory process by augmenting the zymogen activation of MMP-2.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.2
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• pp.785-791
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• 2014

Crude extracts or phytochemicals obtained from some plants have potential anti-cancer properties. Teucrium persicum is an Iranian endemic plant belonging to the Lamiaceae family which has traditionally been used to relieve abdominal pains. However, the anti-cancer properties of this species of the Teucrium genus have not been investigated previously. In this study, we have used a highly invasive prostate cancer cell line, PC-3, which is an appropriate cell system to study anti-tumor properties of plants. A methanolic extract obtained from T persicum potently inhibited viability of PC-3 cells. The viability of SW480 colon and T47D breast cancer cells was also significantly decreased in the presence of the T persicum extract. Flow cytometry suggested that the reduction of cell viability was due to induction of apoptosis. In addition, the results of wound healing and gelatin zymography experiments supported anti-cell invasion activity of T persicum. Interestingly, sublethal concentrations of T persicum extract induced an epithelial-like morphology in a subpopulation of cells with an increase in E-Cadherin and ${\beta}$-Catenin protein levels at the cell membrane. These results strongly suggest that T persicum is a plant with very potent anti-tumor activity.

• Nutritional Sciences
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• v.9 no.3
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• pp.145-151
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• 2006

Matrix metalloproteinases (MMP) play an important role in the extracellular matrix (ECM) degradation undetphysiological and pathological conditions. The present study examined the influence of (-)epigallocatechin gallate and quercetin on phorbol-12-myristate 13-acetate (PMA)-induced secretion of MMP-2 and MMP-9, when human umbilical vein endothelial cells (HUVEC) were treated with (-)epigallocatechin gallate and quercetin at supraphysiological concentrations of $25{\mu}mol/L$. No cytotoxicity was observed by MIT assay in response to a treatment with PMA in the presence of (-)epigallocatechin gallate and quercetin. Western blot analysis and gelatin zymography revealed that exposure of HUVEC to PMA enhanced the levels and gelatinolytic activities of pro and active forms of MMP-2 and active form of MMP-9. (-)Epigallocatechin gallate attenuated PMA-stimulated secretion of active forms of MMP-2 and MMP-9 concomitantly with a loss of activities of these enzymes, which was related to the decreased mRNA levels of MMP. Quercetin was more potent than (-)epigallocatechin gallate in alleviating MMP-9 protein secretion and activity with a decrease in MMP-9 mRNA accumulation. Taken together, the results indicated that (-)epigallocatechin gallte and quercetin exhibited inhibitory effects on MMP activity and may qualify as chemopreventive and cardiovascular protective agents.