Cardioprotective polyphenols from Geum japonicum var. chinense.

Seven undescribed tannins, namely gejaponin A-G, and one dehydrodigallic acid derivative 3,4-dihydroxy-5-(3,4,5-trihydroxy-1-ethoxycarbonyl phenoxy)benzoic acid, together with eighteen known polyphenols were isolated from the 95% ethanol extract of the aerial part of Geum japonicum Thunb. var. chinense F. Bolle. Their structures were elucidated on the basis of comprehensive analysis of UV, IR, NMR, HRMS, and CD spectroscopy experiments. To evaluate their bioactivities, sixteen major compounds were selected to intervene in hydrogen peroxide (H2O2)-induced oxidative damage on H9c2 rat cardiomyoblasts. Some compounds demonstrated high activity in this assay, of which, the known compounds 16 and 21 exhibited strong protective effects against H2O2-induced injury in H9c2 rat cardiomyoblasts, with a comparable cardioprotective activity as that of the positive control trimetazidine, thereby revealing cardioprotective activities from G. japonicum var. chinense.

Evaluation of Geum urbanum L. Extracts with Respect to Their Antimicrobial Potential.

Preparations derived from roots and rhizomes of Geum urbanum L. are traditionally used for the treatment of ulcers and irritations of mucous membranes of the mouth, stomach, and intestinal tract. In complementary medicine, fermentation is one of the methods applied to recover plant extracts used for the production of such pharmaceutical preparations. The present study was performed to characterize the secondary metabolites and to evaluate the antimicrobial potential of different G. urbanum root and rhizome extracts. For this purpose, individual metabolites of fresh and fermented G. urbanum root and rhizome extracts were analyzed by HPLC-DAD-MSn and GC/MS. Among others, rare ellagitannin-sulfates could be characterized by LC/MSn . In addition, the antibacterial activity of various extracts of fresh and dried G. urbanum roots and rhizomes against Staphylococcus aureus (ATCC 6538) and Cutibacterium acnes (CP033842.1; FDAARGOS 503 chromosome) were assessed and compared to that of G. rivale. Furthermore, low- and high-molecular tannins were fractionated by column chromatography, demonstrating the latter to exhibit highest antibacterial activity.

In Vitro Antineoplastic and Antiviral Activity and In Vivo Toxicity of Geum urbanum L. Extracts.

This study evaluated the in vitro antineoplastic and antiviral potential and in vivo toxicity of twelve extracts with different polarity obtained from the herbaceous perennial plant Geum urbanum L. (Rosaceae). In vitro cytotoxicity was determined by ISO 10993-5/2009 on bladder cancer, (T-24 and BC-3C), liver carcinoma (HEP-G2) and normal embryonic kidney (HEK-293) cell lines. The antineoplastic activity was elucidated through assays of cell clonogenicity, apoptosis induction, nuclear factor kappa B p65 (NFκB p65) activation and total glutathione levels. Neutral red uptake study was applied for antiviral activity. The most promising G. urbanum extract was analyzed by UHPLC-HRMS. The acute in vivo toxicity analysis was carried out following OEDC 423. The ethyl acetate extract of aerial parts (EtOAc-AP) exhibited the strongest antineoplastic activity on bladder cancer cell lines (IC50 = 21.33-25.28 µg/mL) by inducing apoptosis and inhibiting NFκB p65 and cell clonogenicity. EtOAc and n-butanol extracts showed moderate antiviral activity against human adenovirus type 5 and human simplex virus type I. Seventy four secondary metabolites (gallic and ellagic acid derivatives, phenolic acids, flavonoids, etc.) were identified in EtOAc-AP by UHPLC-HRMS. This extract induced no signs of acute toxicity in liver and kidney specimens of H-albino mice in doses up to 210 mg/kg. In conclusion, our study contributes substantially to the detailed pharmacological characterization of G. urbanum, thus helping the development of health-promoting phytopreparations.

Protection of the Geum japonicum Thunb. var. chinense extracts against oxygen-glucose deprivation and re-oxygenation induced astrocytes injury via BDNF/PI3K/Akt/CREB pathway.

Geum japonicum Tunb. var. chinense (GJ) is a traditional Chinese medicine usually used for the alleviation of dizziness and headache. Previous studies have reported that the GJ extracts could alleviate cerebral I/R injury by reducing apoptosis in vivo. To further elucidate the positive role and underlying mechanism of the GJ extracts in cerebral I/R injury, the current study investigated the effects of the GJ extracts on oxygen-glucose deprivation and re-oxygenation (OGD/R)-induced astrocytes injury in light of BDNF/PI3K/Akt/CREB signaling pathway with seropharmacological method. In the present study, the LC-MS profiling of the GJ extracts, obtain by reflux extraction, led to the identification of three possible active components were 5-desgalloylstachyurin, tellimagrandin II (TG II) and 3,4,5-Trihydroxybenzaldehyde (THBA). Drug-containing serum was collected from rats given different doses of the GJ extracts (0, 1.75 g/kg, 7 g/kg). Data indicated that the GJ extracts could increase the cell viability and decrease apoptosis and the expression of glial fibrillary acidic protein (GFAP) in OGD/R-induced astrocytes. In addition, the detection of apoptosis-related factors showed that the GJ extracts could obviously increase the expression of Bcl-2 and reduce the expression of Bax, Caspase-3 and cleaved-Caspase-3. Furthermore, the GJ extracts markedly increased the expression of BDNF, TrkB, PI3K, p-Akt and p-CREB. All these effects of the GJ extracts could be significantly reversed by LY294002, an inhibitor of PI3K. These data indicated that the GJ extracts could protect astrocytes against OGD/R-induced injury by inhibiting astrocytes reactivity and apoptosis, owing to the activation of the BDNF/PI3K/Akt/CREB pathway. The results support the application of the GJ extracts in the treatment of ischemic stroke and other ischemic encephalopathy.

Multimedia distributions, bioaccumulation, and trophic transfer of microcystins in the Geum River Estuary, Korea: Application of compound-specific isotope analysis of amino acids.

To determine distributions, bioaccumulation, and trophic transfer of freshwater cyanobacterial toxins such as microcystins (MCs), surface water, suspended solids, sediments, and coastal organisms were collected from seven stations in inner and outer regions of the estuary dam in the Geum River Estuary in June and July 2017. Concentrations of MC variants (MC-LR, -RR, and -YR) in the multimedia samples were analyzed using a HPLC-MS/MS. Trophic position (TP) of organisms (fish, bivalve, gastropod, decapod, and polychaete) was determined by nitrogen stable isotope analyses of both bulk tissues and amino acids. From July to August 2017, great concentrations of MCs were detected in discharged freshwater ranging from 0.4 to 75 µg L-1. Considerable amounts of MCs are delivered to the Geum River Estuary in summer season. MCs spread far away as dissolved phases (18.7-49.5 ng L-1) in July when large amount of freshwater was discharged during the rainy season. Concentrations of MCs in marine organisms varied among species, ranging from 40 to 870 ng g-1 dw. Bioaccumulated MCs tend to decrease with increasing TP of organisms, suggesting that MCs are biodiluted through the marine food web. Compound-specific isotope analysis (nitrogen of amino acids) provides more reliable TPs compared with those by bulk isotope analysis in a closed estuary (such as the Geum River Estuary) with large fluctuations in the isotope ratio of primary producers.

Administration of Asian Herb Bennet (Geum japonicum) Extract Reverses Depressive-Like Behaviors in Mouse Model of Depression Induced by Corticosterone.

Geum japonicum, commonly known as Asian herb bennet, has been used as a diuretic, astringent, anti-dizziness, and anti-headache agent in traditional medicine. Since the antidepressant-like effects of G. japonicum extract have not been well studied, we examined the antidepressant-like effects of G. japonicum extract using depressive-like behavior induced in mice through daily injection of corticosterone (CORT). ICR mice (male, 8 weeks old) were treated with CORT (40 mg/kg, i.p.) and orally administered using oral gavage needles with G. japonicum extract (30, 100, and 300 mg/kg) for 4 weeks. Behavioral experiments were performed 1 h after administration. The control mice exhibited a significant increase in the immobility times in the tail suspension and forced swim tests as well as the step-through latency time in the passive avoidance test. Further, the control group showed a significant decrease in their sucrose consumption. However, treatment with G. japonicum extract at doses of 100 and 300 mg/kg significantly improved these depression-like behaviors without altering the locomotor activity. Moreover, treatment with G. japonicum extract significantly prevented the decrease in the expression of brain-derived neurotrophic factor (BDNF) in the hippocampus. In addition, G. japonicum extract had neuroprotective effects against CORT-induced neurotoxicity in SH-SY5Y cells. Our study indicates that G. japonicum extract exhibits antidepressant-like activity in CORT-induced depressive mice, which might be as a result of increased BDNF expression.

Cardiogenic effects of characterized Geum urbanum extracts on adipose-derived human mesenchymal stem cells.

Stem cell therapy is considered as a promising treatment for cardiovascular diseases. Adipose-derived mesenchymal stem cells (ADMSCs) have the ability to undergo cardiomyogenesis. Medicinal plants are effective and safe candidates for cell differentiation. Therefore, the aim of our study was to investigate cardiogenic effects of characterized (HPLC-UV) extracts of Geum urbanum on ADMSCs of adipose tissue. The methanolic extracts of the root and aerial parts of G. urbanum were obtained and MTT assay was used for studying their cytotoxic effects. Then, cells were treated with 50 or 100 µg/mL of the extracts from root and aerial parts of G. urbanum. MTT assay showed that the extracts of G. urbanum did not have any toxic effects on ADMSCs. Immunostaining results showed increase in the expression of α-actinin and cardiac troponin I (cTnI), and quantitative real-time reverse-transcription PCR data confirmed the upregulation of ACTN, ACTC1, and TNNI3 genes in ADMSCs after treatment. According to HPLC fingerprinting, some cardiogenic effects of G. urbanum extracts are probably due to ellagic and gallic acid derivatives. Our findings indicated that G. urbanum extracts effectively upregulated some essential cardiogenic markers, which confirmed the therapeutic role of this plant as a traditional cardiac medicine.

Chemical constituents of Geum urbanum L. roots.

A new dehydrodigallic acid derivative (1), along with 19 known compounds, including 6 phenolic derivatives, 2 steroids and 11 triterpenoids were isolated from the ethanol extract of the root of Geum urbanum. Herein, there is the first report of steroid and triterpene in Geum urbanum. The purified metabolites were characterised by NMR spectroscopic and mass spectrometric analyses. The identification of the known compounds (2-20) was based on the comparison of their NMR spectroscopic features with previously published data. The structural characteristics of compound 1 were elucidated by comprehensive 1D and 2D NMR spectroscopic methods and acid hydrolysis.

Niga-ichigoside F1 ameliorates high-fat diet-induced hepatic steatosis in male mice by Nrf2 activation.

Hepatic lipid accumulation and oxidative stress (OS) lead to non-alcoholic fatty liver disease (NAFLD). Thus, we hypothesized that antihyperlipidemic and antioxidant activities of niga-ichigoside F1 (NI) would ameliorate events leading to NAFLD. Lanbuzheng (Geum japonicum Thunb. var. chinense), a type of wild vegetable found in Southwest China, was used to extract NI. Male C57BL/6J mice were fed a standard diet (Con) or a high-fat diet (HFD) (denoted as diet) with or without 40 mg kg-1 NI (defined as treatment) for 12 weeks. Diet-treatment interactions were observed in the final body weight, fat pad mass, respiratory exchange ratio (RER) in the daytime, and energy expenditure during the whole day. Moreover, NI alleviated hepatic steatosis, possibly by significantly interacting with HFD to regulate lipid metabolism genes (including Srebp1c, Acc1, Fasn, Scd1, Cpt1a and Fabp5). We also found significant diet-treatment interactions on superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) activities, and thiobarbituric acid reactive substance (TBARS) levels, as well as the nuclear and cellular Nrf2 protein levels. Significant free fatty acid (FFA)-treatment interactions on Nrf2 nuclear translocation, antioxidant enzymes activities, genes in lipogenesis (Srebp1c, Acc1, Fasn, and Scd1), and fatty acid oxidation (Pparα) and transport (Fabp5 and Cd36) were also detected in 1 mM FFA-treated HepG2 cells with or without 20 µM NI. These beneficial effects of NI on oxidative stress and lipid accumulation were abolished by Nrf2 siRNA. Our data revealed that dietary NI could prevent HFD-induced hepatic steatosis, possibly via interacting with HFD to activate Nrf2 nuclear translocation to maintain a redox status, thus regulating lipid metabolism genes expressions.

Mechanistic study of the inhibitory activity of Geum urbanum extract against α-Synuclein fibrillation.

The presence of Lewy bodies and Lewy neurites is a major pathological hallmark of Parkinson's disease and is hypothesized to be linked to disease development, although this is not yet conclusive. Lewy bodies and Lewy neurites primarily consist of fibrillated α-Synuclein; yet, there is no treatment available targeting stabilization of α-Synuclein in its native state. The aim of the present study was to investigate the inhibitory activity of an ethanolic extract of Geum urbanum against α-Synuclein fibrillation and examine the structural changes of α-Synuclein in the presence of the extract. The anti-fibrillation and anti-aggregation activities of the plant extract were monitored by thioflavin T fibrillation assays and size exclusion chromatography, while structural changes were followed by circular dichroism, Fourier transform infrared spectroscopy, intrinsic fluorescence, small angle X-ray scattering and electron microscopy. Since the extract is a complex mixture, structure-function relationships could not be determined. Under the experimental conditions investigated, Geum urbanum was found to inhibit α-Synuclein fibrillation in a concentration dependent way, and to partly disintegrate preformed α-Synuclein fibrils. Based on the structural changes of α-Synuclein in the presence of extract, we propose that Geum urbanum delays α-Synuclein fibrillation either by reducing the fibrillation ability of one or more of the aggregation prone intermediates or by directing α-Synuclein aggregation towards a non-fibrillar state. However, whether these alterations of the fibrillation pathway lead to less pathogenic species is yet to be determined.

Effects of Geum urbanum L. root extracts and its constituents on polymorphonuclear leucocytes functions. Significance in periodontal diseases.

ETHNOPHARMACOLOGICAL RELEVANCE: Geum urbanum L. (wood avens) root infusions and decoctions have been used externally for reducing the bleeding and inflammation of gums (gingivitis), and mucous membranes. AIM OF THE STUDY: Taking into account that primed and hyperactivated neutrophils are an important factor in the transition from gingivitis to periodontitis, we investigated the effects of phytochemically characterised (HPLC-DAD-MS(n)) extracts of different polarity from Geum urbanum root on oxidative burst, elastase, metalloproteinase 9 (MMP-9), interleukin 8 (IL-8) and 1ß (IL-1ß), tumour necrosis factor (TNF-α) release, expression of adhesion molecules (CD62L and CD11b) and delayed apoptosis in stimulated neutrophils. As gemin A is a dominating compound in a raw material, so we considered its activity in parallel with the positive control quercetin. MATERIALS AND METHODS: The extracts were characterised by HPLC-DAD- MS(n) method. The inhibition of ROS production by stimulated neutrophils was determined using luminol dependent chemiluminescence method. The effect on MMP-9, IL-1ß, TNF-α and IL-8 production by neutrophils was measured by enzyme-linked immunosorbent assay (ELISA). Neutrophil elastase release was established spectrophotometrically. The expression of adhesion molecules and the apoptosis of neutrophils was analyzed with flow cytometry. RESULTS: The main compounds detected in the extract belong mainly to the group of ellagitannin: pedunculagin, stachyurin, casuarynin and gemin A, and ellagic acid derivatives. Procyanidins and one complex tannin were found as minor compounds. Gemin A significantly affected the functions of stimulated neutrophils by reducing the surface expression of CD11b, and inhibiting the release of reactive oxygen species, and proteases (elastase, MMP-9), chemokines and cytokines (interleukins IL-8, IL-1ß). Interestingly, gemin A stimulated the release of TNF-α, which may be one of the stimulators of apoptosis of neutrophil cells. The primary aqueous extract, the ethyl acetate and the butanolic fractions, all containing the highest level of gemin A, have exerted similar but weaker activity. CONCLUSION: The modulating effect on the neutrophils function of extracts, and its main constituent gemin A, support the traditional use of this plant material in cavity inflammation including mucositis, gingivitis and periodontosis.

Composition of essential oil from aerial and underground parts of Geum rivale and G. urbanum growing in Poland.

A study of the composition of essential oils from aerial and underground parts of Geum rivale L. and Geum urbanum L. growing in Poland led to the identification of 130 compounds. The main compound of the essential oil from underground parts of G. urbanum was eugenol (69.2%), whereas cis-myrtanal (53.3%) was the major constituent of the essential oil from roots of G. rivale. The essential oils from aerial parts of the plants contained large amounts of aliphatic compounds with (Z)-3-hexenol (38.4%) being the dominant constituent of the essential oil from aerial parts of G. urbanum and 1-octen-3-ol (33.9%) from G. rivale.

Investigation into biologically active constituents of Geum rivale L.

Aerial and underground parts of Geum rivale (Rosaceae) were investigated. Tiliroside, gallic acid, ellagic acid and a sterol fraction were isolated from aerial parts of the plant. The sterol fraction was analyzed using GC-MS. Eleven phenolic acids were identified in aerial parts of the plant, and eight in underground parts, by means of RP-HPLC analysis. The quantitative determination of phenolic acids, tannins and flavonoids was also carried out.

Anti-Helicobacter pylori activity of the methanolic extract of Geum iranicum and its main compounds.

Geum iranicum Khatamsaz, belonging to the Rosaceae family, is an endemic plant of Iran. The methanol extract of the roots of this plant showed significant activity against one of the clinical isolates of Helicobacter pylori which was resistant to metronidazole. The aim of this study was the isolation and evaluation of the major compounds of G. iranicum effective against H. pylori. The compounds were isolated using various chromatographic methods and identified by spectroscopic data (1H and 13C NMR, HMQC, HMBC, EI-MS). An antimicrobial susceptibility test was performed employing the disk diffusion method against clinical isolates of H. pylori and a micro dilution method against several Gram-positive and Gram-negative bacteria; additionally the inhibition zone diameters (IZD) and minimum inhibitory concentrations (MIC) values were recorded. Nine compounds were isolated: two triterpenoids, uvaol and niga-ichigoside F1, three sterols, beta-sitosterol, beta-sitosteryl acetate, and beta-sitosteryl linoleate, one phenyl propanoid, eugenol, one phenolic glycoside, gein, one flavanol, (+)-catechin, and sucrose. The aqueous fraction, obtained by partitioning the MeOH extract with water and chloroform, was the most effective fraction of the extract against all clinical isolates of H. pylori. Further investigation of the isolated compounds showed that eugenol was effective against H. pylori but gein, diglycosidic eugenol, did not exhibit any activity against H. pylori. The subfraction D4 was the effective fraction which contained tannins. It appeared that tannins were probably the active compounds responsible for the anti-H. pylori activity of G. iranicum. The aqueous fraction showed a moderate inhibitory activity against both Gram-positive and Gram-negative bacteria. The MIC values indicated that Gram-positive bacteria including Staphylococcus aureus, Staphylococcus epidermidis, and Bacillus subtilis are more susceptible than Gram-neagative bacteria including Escherichia coli and Pseudomonas aeruginosa.

A natural compound induced cardiogenic differentiation of endogenous MSCs for repair of infarcted heart.

An intra-myocardial injection of a cardiogenic factor (cardiogenin) was reported to induce myocardial regeneration of exogenous mesenchymal stem cell (MSCs) origin. In this study, replacement of the dangerous intra-myocardial injection with a safe method and whether the endogenous MSCs contribute to the cardiogenin-mediated myocardial regeneration were investigated. Bone marrow transplantation with labeled MSCs was performed in rats, which were subsequently subject to a permanent ligation of left anterior descending coronary artery one week after the transplantation. The rats were then treated with the cardiogenin through oral administration for 2 weeks. We not only demonstrated the substantial therapeutic effects of cardiogenin on myocardial infarction through an oral administration, but also provided direct evidences that the bone marrow derived endogenous MSCs are the major cellular source of the regenerating myocardium. Preliminary mechanistic studies suggested that miR-9 and its target E-cadherin may be required for intercalated disc formation.

[Myocardial regeneration and repair of infarcted heart by a new composition isolated from Geum japonicum].

OBJECTIVES: To isolate the cardiogenic fraction, which can enhance cardiogenic differentiation of bone marrow-derived mesenchymal stem cells (MSC) from Geum japonicum. The therapeutic effect of the isolated cardiogenic fraction was further tested in a rat myocardial infarction (MI) model. METHOD: Bioassay guided fractionation method was used for the isolation of the cardiogenic fraction, named as heart repair fraction (HRF). MI was induced by a permanent ligation of left anterior descending coronary artery. The rats exhibiting similarly decreased values of left ventricle ejection fraction (LVEF) and fraction shortening (LVFS) were used. The rats in test group (n = 10) were subject to HRF treatment (20 mg×kg(-1)×d(-1)) through gastric gavage daily for 4 weeks. Water alone (2 ml/d) was given through gastric gavage to rats in the control group (n = 10). The cardiac function was assessed by echocardiography at different time points. Masson trichrome staining was used for evaluation of the infarct size. Morphological and immunohistochemical studies were performed to investigate the HRF mediated myocardial regeneration. RESULTS: LVEF (66.2% ± 6.9%) and LVFS (46.8% ± 5.8%) were significantly increased two weeks post HRF treatment compared with the values (LVEF: 55.7% ± 6.0% and LVFS: 36.4% ± 5.2%) in control rats (all P < 0.01). The improved heart function was further restored 4 weeks post HRF treatment (P < 0.01). Furthermore, the treatment of acute MI with this HRF significantly reduced the infarct size (19.0% ± 6.1%) compared with that (31.1% ± 8.6%) in control rats (P < 0.01). Substantial regeneration of cardiomyocytes in infarcted region of the HRF treated heart was also observed that replaced a considerable part of the infarcted heart tissues resulting in remarkable reduction of the infarct size. CONCLUSION: The properties of this HRF isolated from Geum japonicum in stimulating substantial regeneration of myocardium in infarct region with consequently improved cardiac function appear to be new and represent a new approach for the treatment of MI.

Taraxasterane-type triterpene and neolignans from Geum japonicum Thunb. var. chinense F. Bolle.

The phytochemical investigation of the ethanolic extract of the whole plants of Geum japonicum Thunb. var. chinense F. Bolle yielded four new compounds: 20 ß,28-epoxy-28-hydroxytaraxasteran-3 ß-ol (1), (7R,8R)-4-hydroxy-9'-O-(α-L-rhamnopyranosyl)-3,3',5'-trimethoxy-8-O-4'-neolignan (2), (7R,8S)-4-hydroxy-9'-O-(α-L-rhamnopyranosyl)-3,3',5'-trimethoxy-8-O-4'-neolignan (3), and (7S,8S)-5-methoxycupressoside A (4), as well as 40 other known compounds. Structures were elucidated by physical, chemical, and spectroscopic methods, including 1D and 2D NMR, HRESIMS, and CD experiments. The taraxasterane-type triterpene (1) and lignans (2- 6) are reported for the first time from the GEUM genus. Moreover, all compounds were evaluated for anti-inflammatory activities against NO production in RAW264.7 macrophages, and only moderate activities were detected.

Fatty acid synthase inhibitors from Geum japonicum Thunb. var. chinense.

Bioassay-guided fractionation of the MeOH extract of Geum japonicum Thunb. var. chinense using the fatty acid synthase inhibition assay led to the isolation of a new dimeric ellagitannin, gemin G (1), together with six known compounds, gemin A (2), casuarinin (3), pedunculagin (4), potentillin (5) , tellimagrandin II (6), and ellagic acid (7). Their structures were determined on the basis of spectroscopic analyses. Compounds 1-7 displayed strong inhibitory activities on fatty acid synthase with IC(50) values in the range of 0.21-41.4 microM. Compounds 1-4 exhibited significant antioxidant activities higher than vitamin C in the ORAC assay. Compounds 1 and 2 also showed weak cytotoxic effects on BGC-823 cell.

3,4,5-trihydroxybenzaldehyde from Geum japonicum has dual inhibitory effect on matrix metalloproteinase 9; inhibition of gelatinoytic activity as well as MMP-9 expression in TNF-alpha induced HASMC.

The matrix metalloproteinases (MMP-9 and MMP-2) production and smooth muscle cell (SMC) migration may play key roles in the pathogenesis of atherosclerotic lesions. In particular, the cancer cell invasion and SMC migration through vascular wall were shown to be directly associated with inducible MMP-9 expression. Previously, 3,4,5-trihydoroxybenzaldehyde (THBA) was purified from Geum japonicum and we demonstrated a direct inhibition effect of THBA on MMP-9 and MMP-2 activity in the supernatants of TNF-alpha-induced HASMCs. In addition, MMP-9 expression and migration was suppressed by THBA in the TNF-alpha-induced HASMCs. In this study, we also investigated whether TNF-alpha-induced MMP-9 expressions are involved with migrations of HASMCs by using cell signal inhibitors and MMP-9 inhibitors. An RT-PCR and luciferase-tagged promoter analysis revealed that THBA inhibits the transcription of MMP-9 mRNA. Moreover, an electrophoretic mobility shift assay (EMSA) exhibited that THBA also suppressed DNA binding of nuclear factor (NF)-kappaB and activator protein (AP)-1 transcription factors. Furthermore, Western blot analysis indicated TNF-alpha-induced phosphorylation of extracellular signal regulated kinase 1 and 2 (ERK1/2), p38 and c-Jun N-terminal kinase (JNK) were inhibited by THBA. Taken together, we suggest that THBA has inhibition effect to the migrations as well as MMP-2 and MMP-9 activities in HASMCs. Especially gelatinolytic activity was controlled by enzymatic inhibition of MMP-2 and MMP-9, and also down-regulated MMP-9 transcription via mitogen-activated protein kinase (MAPK) pathways in THBA treated HASMCs.