• Journal of Life Science
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• v.26 no.5
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• pp.614-619
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• 2016

The prostate-apoptosis-response-gene-4 (Par-4) protein has been identified as an effector of cell death in response to various apoptotic stimuli in prostate cancer cells. We found that overexpression of Par-4 by stable transfection inhibits cell migration and invasion in Caki cells. The expression of various matrix metalloproteinases (MMPs) has been implicated in the invasion and metastasis of cancer cells. In this study, we investigated whether ectopic expression of Par-4 modulates MMP-2 expression and activity in human renal carcinoma Caki cells. We found that overexpression of Par-4 markedly inhibited MMP-2 activity, but not MMP-9 activity. However, loss of the leucine zipper domain of Par-4 (Par-4 ΔLZ#1 and #2) did not inhibit MMP-2 activity. Further, knock-down of Par-4 with the corresponding siRNA resulted in increased invasion and metastasis of renal carcinoma Caki cells. Interestingly, overexpression or knock-down of Par-4 did not affect the expression levels of MMP-2 mRNA. Taken together, our findings suggest that Par-4 may inhibit MMP-2 activity through its post-transcriptional regulation in renal carcinoma Caki cells.

• Journal of Periodontal and Implant Science
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• v.43 no.1
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• pp.24-29
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• 2013

Purpose: Matrix metalloproteinases (MMPs) are capable of degrading extracellular matrix, and they are inducible enzymes depending on an inflammatory environment such as periodontitis and bacterial infection in periodontal tissue. Gingival inflammation has been postulated to be correlated with the production of MMP-2 and MMP-9. The objective of this study was to quantify the expression and activity of MMP-9 and -2, and to determine the correlation between activity and expression of these MMPs in human gingival tissues with periodontitis. Methods: The gingival tissues of 13 patients were homogenized in $500{\mu}L$ of phosphate buffered saline with a protease inhibitor cocktail. The expression and activity of MMP-2 and -9 were measured by enzyme-linked immunosorbent assay and Western blot analysis, and quantified by a densitometer. For the correlation line, statistical analysis was performed using the Systat software package. Results: MMP-9 was highly expressed in all gingival tissue samples, whereas MMP-2 was underexpressed compared with MMP-9. MMP-9 activity increased together with the MMP-9 expression level, with a positive correlation (r=0.793, P=0.01). The correlation was not observed in MMP-2. Conclusions: The expression of MMP-2 and -9 might contribute to periodontal physiological and pathological processes, and the degree of MMP-9 expression and activity are predictive indicators relevant to the progression of periodontitis.

• Applied Biological Chemistry
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• v.46 no.1
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• pp.60-65
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• 2003

MMP-1 inhibitory compounds were isolated from 120 Korean traditional edible plants. UP- 1 activity significantly increased linearly with increasing UVB dose in normal human foreskin fibroblast HS68 cell, showing maximum activity at approximately 35 $mJ/cm^2$, whereas in HaCaT cell, normal human keratinocyte, no increase was observed. Maximum secretion of MMP-1 after UVB treatment occurred around 36-48 k after treatment. MMP-1 inhibitory compound isolated from cold-water fraction of Cataegus pinnatifida Bunge showed the mort potent activity. The MMP-1 inhibitory compound was deduced as a peptide based on the fact that pronase digestion decreased the activity whereas periodate oxidation did not. The most potent UP- 1-inhibitory protein, CP-2Va-2, showing an activity of 88.5% against MMP-1, was isolated through sequential column chromatography on DEAE-Toyopearl 650C, Butyl-Toyopearl 650M, and Bio-Gel P-30. Molecular weight of CP-2Va-2 determined through high performance liquid chromatography and SDS PACE was 19 and 20 kDa. respectively, signifying a monomeric structure.

• Environmental Analysis Health and Toxicology
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• v.24 no.1
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• pp.63-70
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• 2009

Naringenin, major one of the citrus flavonoids, have been identified that exert antioxidative, anticancer effects. The present study investigated the effects of naringenin on tumor invasion and matrix metalloproteinases(MMPs) activities. Naringenin inhibited cell invasion of HT-1080 fibrosarcoma cells in a dose-dependent manner. The activities of MMP-2 and MMP-9 were inhibited by naringenin as demonstrated by gelatin zymography assay. Furthermore, the amounts of MMP-2, MMP-9, and MT1-MMP mRNA were analyzed in the cells. MMP-2, MMP-9, and MT1-MMP mRNA expression were suppressed by naringenin with time and dose-dependent. These results demonstrate that anti-metastatic activities of naringenin resulted from blocking of invasion of the HT-1080 cells. Taken together, the results of this studies provide evidence that naringenin possess an anti-metastatic activity.

• The Korean Journal of Physiology and Pharmacology
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• v.20 no.2
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• pp.221-228
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• 2016

We investigated whether prunetin affects the proteolytic activity, secretion, and gene expression of matrix metalloproteinase-3 (MMP-3) in primary cultured rabbit articular chondrocytes, as well as in vivo production of MMP-3 in the rat knee joint to evaluate the potential chondroprotective effect of prunetin. Rabbit articular chondrocytes were cultured in a monolayer, and reverse transcriptionpolymerase chain reaction (RT-PCR) was used to measure interleukin-$1{\beta}$ (IL-$1{\beta}$)-induced expression of MMP-3, MMP-1, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4), and ADAMTS-5. In rabbit articular chondrocytes, the effects of prunetin on IL-$1{\beta}$-induced secretion and proteolytic activity of MMP-3 were investigated using western blot analysis and casein zymography, respectively. The effect of prunetin on MMP-3 protein production was also examined in vivo. The results were as follows: (1) prunetin inhibited the gene expression of MMP-3, MMP-1, MMP-13, ADAMTS-4, and ADAMTS-5; (2) prunetin inhibited the secretion and proteolytic activity of MMP-3; (3) prunetin suppressed the production of MMP-3 protein in vivo. These results suggest that prunetin can regulate the gene expression, secretion, and proteolytic activity of MMP-3, by directly acting on articular chondrocytes.

• Journal of Life Science
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• v.26 no.2
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• pp.174-180
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• 2016

Dicumarol is a coumarin derivative isolated from sweet clover (Melilotus alba), and has anti-coagulant activity with the inhibitory activity of NAD(P)H quinone oxidoreductase1 (NQO1). NQO1 catalyzes the two-electron reduction of quinones to hydroquinones. Dicumarol competes with NAD(P)H for binding to NQO1, resulting in the inhibition of NQO1 enzymatic activity. The expression of matrix metalloproteinases (MMPs) has been implicated in the invasion and metastasis of cancer cells. The expression of MMPs is regulated by cytokines and signal transduction pathways, including those activated by phorbol myristate acetate (PMA). However, the effects of dicumarol on metalloproteinase (MMP)-9 expression and activity are not investigated here. This study investigated whether dicumarol inhibits MMP-9 expression and activity in PMA-treated human renal carcinoma Caki cells. Dicumarol markedly inhibited the PMA-induced MMP-9 mRNA expression and MMP-9 activity. NF-κB and AP1 promoter activity, which is important in MMP-9 expression, also decreased in dicumarol-treated cells. Furthermore, dicumarol markedly suppressed the ability of PMA-mediated migration in Caki cells. When the relevance of NQO1 in the dicumarol-mediated inhibitory effect on PMA-induced MMP9 activity was elucidated, knock-down of NQO1 with siRNA was found to have no effect on PMA-induced MMP9 activity, suggesting that the stimulating effect of dicumarol on PMA-induced MMP9 activity is independent of NQO1 activity. Taken together, the present studies suggested that dicumarol may inhibit PMA-induced migration via down-regulation of MMP-9 expression and activity.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.33 no.6
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• pp.636-642
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• 2007

Background: CpG DNA plays an important role in immune cell function. This study examined whether the temporal control of toll-like receptor (TLR)9 by CpG DNA can regulate the expression of matrix metalloproteinase-9(MMP-9). Methods and materials: Macrophages were cultured in the presence of 10% FBS. For the various MMP genes analysis, RT-PCR and real-time PCR were performed. In addition, zymography assay performed for the MMP activity. The phosphorylation assay did for the ERK1/2 and NF${\kappa}B$ activation, and luciferase promoter assay was for the NF${\kappa}B$ activity. Results: CpG DNA induced the mRNA expression of MMP-2, MMP-9, and MMP-13, but not of MMP-7, MMP-8, and MMP-12, in a time-dependent manner. Especially, the mRNA expression of MMP-9 was strongly induced by CpG DNA using real-time RT-PCR. The TLR9 inhibitor, chloroquine, suppressed CpG DNA-induced MMP-9 expression and its activity. Moreover, CpG DNA induced the phosphorylation of ERK and the inhibition of ERK by U0126 suppressed CpG DNA-induced MMP-9 expression and its activity. CpG DNA stimulated $I{\kappa}B-{\alpha}$ degradation and luciferase activity. In addition, pretreatment of SN-50, the inhibitor of NF${\kappa}B$, strongly blocked the CpG DNA-induced MMP-9 expression and activity. Conclusion: These observations suggest that CpG DNA may play important roles in the activation of macrophages by regulating the production of MMP-9 via the sequential TLR9-ERK-NF${\kappa}B$ signaling pathway.

• Journal of Life Science
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• v.26 no.5
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• pp.584-591
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• 2016

Saururus chinensis is a perennial plants, its flavonoid compound is known to exhibit anti-oxidative activity. This study was aimed to investigate the effect of Water Extract and Solvent Fractions of Saururus chinensis on antioxidant, anti-inflammatory and anti-invasive of Phorbol 12-myristate 13-acetate (PMA)-induced matrix metalloproteinase (MMP-2) and MMP-9 activities. Plant samples were fractionated into hexane, CHCl₃, ethyl acetate, butanol, and water fractions, and each of these was assayed individually. The water fraction showed the highest extraction yield at 9.25%(w/w). Anti-oxidative activity was analyzed by DPPH assay. Cell viability was detected by the MTS assay. Anti-inflammatory activity was assayed by the nitric oxide (NO) production in mouse macrophage Raw 264.7 cells. The activity and mRNA expression of MMP-2 and MMP-9 in human oral squamous carcinoma YD-10B cells were examined by zymography and RT-PCR. As results, MMP-2/-9 activation was increased in PMA induced YD-10B cells. In PMA-treated YD-10B cells, the increased mRNA expression and protein activation of MMP-2/-9 were significantly inhibited in the ethyl acetate fraction. The ethyl acetate fraction showed the highest anti-oxidative activity at 73.38%. The ethyl acetate fraction at non-cytotoxic concentrations significantly exhibited the anti-inflammatory activity of Raw 264.7 cells in dose-dependent manner. In conclusion, these findings demonstrate that the ethyl acetate fraction obtained from a chinensis water extract potentiates a promising therapeutic anti-invasive agent and, therefore, as an anti-cancer drug for cancer prevention and therapy in oral cancer.

• Biomolecules & Therapeutics
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• v.10 no.4
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• pp.240-245
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• 2002

Cancer metastasis represents the most important cause of cancer death and antitumor agents that may inhibit this process have been extensively pursued. Invasion and metastasis of malignantly transformed cells involve degradation of the extracellular matrix (ECM) components by matrix metalloproteinases (MMP), especially MMP-2 and -9. We previously showed that H-ras-induced invasive phenotype may involve MMP-2, rather than MMP-9, in MCF10A cells. In the present study, we investigated the chemopreventive effect of Aster, a widely used culinary vegetable in Korea. We screened twelve extracts from three Aster species (Aster scaber, Aster oharai and Aster glehni) for the inhibitory effect on MMP activities of H-ras MCF10A human breast epithelial cells. All of the extracts tested in this study efficiently inhibited the gelatinolytic activities of MMP-2 and MMP-9. A more prominent inhibition was observed in MMP-2 activity compared to MMP-9. Out of twelve extracts, eight extracts showed>90％ inhibition of MMP-2 activity in H-ras MCF10A cells while only one extract showed>90％ inhibition of MMP-9 activity. We selected three extracts (AO-3, AG-3 and AS-EA) for further studies since they exerted a marked inhibition in the ratio of MMP-2 to MMP-9. Treatment with AO-3, AG-3 and AS-EA in H-ras MCF10A cells caused a significant inhibition of invasive phenotype and migration, proving a chemopreventive potential of these extracts. Taken together, our results demonstrate that extracts of Aster effectively inhibit invasion and migration of highly malignant human breast cells, possibly via downregulation of MMP-2 and MMP-9.

• Biomolecules & Therapeutics
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• v.22 no.5
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• pp.414-419
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• 2014

Matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases that regulate cell-matrix composition and are also involved in processing various bioactive molecules such as cell-surface receptors, chemokines, and cytokines. Our group recently reported that MMP-3, -8, and -9 are upregulated during microglial activation and play a role as proinflammatory mediators (Lee et al., 2010, 2014). In particular, we demonstrated that MMP-8 has tumor necrosis factor alpha (TNF-${\alpha}$)-converting enzyme (TACE) activity by cleaving the prodomain of TNF-${\alpha}$ and that inhibition of MMP-8 inhibits TACE activity. The present study was undertaken to compare the effect of MMP-8 inhibitor (M8I) with those of inhibitors of other MMPs, such as MMP-3 (NNGH) or MMP-9 (M9I), in their regulation of TNF-${\alpha}$ activity. We found that the MMP inhibitors suppressed TNF-${\alpha}$ secretion from lipopolysaccharide (LPS)-stimulated BV2 microglial cells in an order of efficacy: M8I>NNGH>M9I. In addition, MMP inhibitors suppressed the activity of recombinant TACE protein in the same efficacy order as that of TNF-${\alpha}$ inhibition (M8I>NNGH>M9I), proving a direct correlation between TACE activity and TNF-${\alpha}$ secretion. A subsequent pro-TNF-${\alpha}$ cleavage assay revealed that both MMP-3 and MMP-9 cleave a prodomain of TNF-${\alpha}$, suggesting that MMP-3 and MMP-9 also have TACE activity. However, the number and position of cleavage sites varied between MMP-3, -8, and -9. Collectively, the concurrent inhibition of MMP and TACE by NNGH, M8I, or M9I may contribute to their strong anti-inflammatory and neuroprotective effects.