Panax notoginseng saponins inhibit areca nut extract-induced oral submucous fibrosis in vitro.

BACKGROUND: Oral submucous fibrosis (OSF) is a premalignant and fibrosing disease, which is closely associated with the habit of chewing areca nut. Panax notoginseng Buck F. H. Chen is an often used antifibrotic and antitumor agent. To treat areca nut-induced OSF, we have developed a chewable tablet, in which one of the major medicines is total Panax notoginseng saponins (PNS). In this study, we have investigated the antifibrotic effect and mechanism of PNS on areca nut-induced OSF in vitro. METHODS: Through human procollagen gene promoter luciferase reporter plasmid, hydroxyproline assay, gelatin zymography, qRT-PCR, ELISA, and Western blot, the influences of PNS on areca nut extract (ANE)-induced cell growth, collagen accumulation, procollagen gene transcription, MMP-2/-9 activity, MMP-1/-13 and TIMP-1/-2 expression, cytokine secretion, and the activation of PI3K/AKT, ERK/JNK/p38 MAPK, and TGFß/Smads pathways were detected. RESULTS: Panax notoginseng saponins could inhibit the ANE-induced abnormal growth and collagen accumulation of oral mucosal fibroblasts in a concentration-dependent manner. PNS (25 µg/ml) could significantly inhibit the ANE-induced expression of Col1A1 and Col3A1, augment the ANE-induced decrease of MMP-2/-9 activity, inhibit the ANE-induced increase of TIMP-1/-2 expression, and decrease the ANE-induced transcription and release of CTGF, TGFß1, IL-6, and TNFα. PNS (25 µg/ml) also significantly inhibited the ANE-induced activation of AKT and ERK/JNK/p38 MAPK pathways in oral mucosal fibroblasts and the ANE-induced activation of TGFß/smad pathway in HaCaT cells. CONCLUSION: Panax notoginseng saponins possess excellent anti-OSF activity, and its mechanism may be related to its ability to inhibit the ANE-induced activation of PI3K/AKT, ERK/JNK/p38 MAPK, and TGFß/smad pathways.

[Effect of spearmint oil on lipopolysaccharide induced emphysema-like changes and expression of matrix metalloproteinase-9].

OBJECTIVE: To investigate the effect of spearmint oil on emphysema-like changes and the expression of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta(IL-1beta), matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-9) in lipopolysaccharide (LPS) treated rats. METHOD: Emphysematous changes model was induced by intratracheal instillation of LPS once a week for up to 8 weeks in rats. Rats were divided into control, dexamethasone (0.3 mg x kg(-1)), and spearmint oil (10, 30,100 mg x kg(-1)) groups. Each group was treated with saline, dexamethasone, and spearmint of oil respectively for 4 weeks. Then total and different white blood cell counts in bronchoalveolar lavage fluid(BALF) were carried out. The pathologic changes of lung tissue such as alveolar structure, airway inflammation, and goblet cell metaplasia were observed by HE and AB-PAS staining. Expression of TNF-alpha, IL-1beta, TIMP-1 and MMP-9 were measured. RESULT: Both spearmint and dexamethasone decreased the destruction of pulmonary alveolus. The total and different white blood cell counts in BALF including neutrophile and lymphocyte of spearmint oil 100 mg x kg(-1) and dexamethasone group were significantly reduced, and the goblet cell metaplasia was also inhibited. Dexamethasone had inhibitory effect on the expression of TNF-alpha, IL-1beta, TIMP-1 and MMP-9. Spearmint oil 30, 100 mg x kg(-1) significantly reduced TNF-alpha and IL-1beta respectively. Spearmint oil 10, 30 and 100 mg x kg(-1) had no effect on the expression of TIMP-1, but could decrease the expression of MMP-9 significantly in lung tissues. CONCLUSION: Spearmint oil has protective effect on rats with emphysematous changes, since it improves alveolar destruction, pulmonary inflammation, and goblet cell metaplasia. The mechanism may include reducing TNF-alpha, IL-1beta content and inhibiting overexpression of matrix metalloproteinase-9 in lung tissues.

[Regulation of calculus bovis on the function of mice oral fibroblasts].

To explore the influence of calculus bovis on the function of primary cultured mice oral fibroblasts, we determined the effects of calculus bovis on the fibroblast proliferation, collagen production, matrix metalloproteinases-2, -9 activities and tissue inhibitor of metalloproteinase-1 production by MTT assay, chloramine T method, gelatin zymography and enzyme-linked immunosorbent assays respectively. The results showed that calculus bovis could significantly inhibit the proliferation of fibroblasts and collagen synthesis in a concentration dependent manner, could significantly (P<0.05) suppress matrix metalloproteinases-2 activity and very significantly (P<0.01) inhibit the production of tissue inhibitor of metalloproteinase-1. In conclusion, the major function of calculus bovis in the process of ulcer healing is not to promote tissue regeneration, the mechanism that calculus bovis inhibits collagen synthesis may be partly due to its ability to very significantly (P<0.01) suppress the production of tissue inhibitor of metalloproteinase-1.

Human rheumatoid arthritis tissue production of IL-17A drives matrix and cartilage degradation: synergy with tumour necrosis factor-alpha, Oncostatin M and response to biologic therapies.

INTRODUCTION: The aim of this study was to examine IL-17A in patients, following anti-TNF-alpha therapy and the effect of IL-17A on matrix turnover and cartilage degradation. METHODS: IL-17A expression was examined by ELISA and immunohistology in the rheumatoid arthritis (RA) joints. RA whole synovial tissue explant (RA ST), primary synovial fibroblasts (RASFC), human cartilage and chondrocyte cultures were stimulated with IL-17A +/- TNF-alpha and Oncostatin M (OSM). Matrix metalloproteinase (MMP) and tissue inhibitor (TIMP-1) were assessed by ELISA and zymography. Cartilage proteoglycan release was assessed histologically by Safranin-O staining. Clinical parameters, IL-17A, MMP/TIMP were assessed in patients pre/post biologic therapy. RESULTS: IL-17A levels were higher in RA vs osteoarthritis (OA)/normal joints (P < 0.05). IL-17A up-regulated MMP-1, -2, -9, and -13 in RA ST, RASFC, cartilage and chondrocyte cultures (P < 0.05). In combination with TNF-alpha and OSM, IL-17A shifted the MMP:TIMP-1 ratio in favor of matrix degradation (all P < 0.05). Cartilage proteoglycan depletion in response to IL-17A was mild; however, in combination with TNF-alpha or OSM showed almost complete proteoglycan depletion. Serum IL-17A was detected in 28% of patients commencing biologic therapy. IL-17A negative patients demonstrated reductions post therapy in serum MMP1/TIMP4, MMP3/TIMP1 and MMP3/TIMP4 ratios and an increase in CS846 (all P < 0.05). No significant changes were observed in IL-17A positive patients. CONCLUSIONS: IL-17A is produced locally in the inflamed RA joint. IL-17A promotes matrix turnover and cartilage destruction, especially in the presence of other cytokines, mimicking the joint environment. IL-17A levels are modulated in vivo, following anti-TNF therapy, and may reflect changes in matrix turnover.

The effects of selective COX-2 inhibitor/celecoxib and omega-3 fatty acid on matrix metalloproteinases, TIMP-1, and laminin-5gamma2-chain immunolocalization in experimental periodontitis.

BACKGROUND: Matrix metalloproteinases (MMPs) play important roles in tissue-destruction mechanisms-associated periodontitis. MMP-8 and -13 are the predominant collagenases that are important in the extracellular matrix degradation in periodontal tissues. MMP-14 is a membrane-type MMP, whereas laminin-5 indicates basal membrane modification and epithelial induction. The purpose of the present study was to evaluate the effects of celecoxib and omega-3 fatty acid administration on the gingival tissue expression of MMP-8, -13, and -14, tissue inhibitor of MMP (TIMP)-1, and laminin (Ln)-5gamma2-chain in rat experimental periodontitis induced by Escherichia coli endotoxin (lipopolysaccharide [LPS]). METHODS: Experimental periodontitis was induced in rats by repeated LPS injection. Fifty-one adult male Sprague-Dawley rats were divided into six study groups: saline control, LPS, LPS + celecoxib, LPS + therapeutic omega-3 (TO3), prophylactic omega-3 + LPS + omega-3 (P+TO3), and LPS + celecoxib + omega-3 fatty acid. Celecoxib and omega-3 fatty acid were given as a single agent or as combination therapy for 14 days. On day 15, all rats were sacrificed, and gingival tissues were analyzed immunohistochemically for the expression of MMP-8, -13, and -14, TIMP-1, and Ln-5gamma2-chain. Alveolar bone loss was evaluated morphometrically under a stereomicroscope. Data were tested statistically by Kruskal-Wallis and Mann-Whitney tests and Spearman correlation analysis. RESULTS: Alveolar bone loss was significantly higher in all study groups compared to the saline control group (all P <0.01). MMP-8 expression was significantly higher in the LPS group than in the saline group (P = 0.001). Very low expression of MMP-8 was found in the celecoxib, P+TO3, and combination groups. TO3 increased TIMP-1 expression significantly compared to the LPS group (P <0.05). Individual celecoxib and P+TO3 administration increased MMP-14 significantly compared to saline control and LPS groups (P <0.05). No significant differences were found among the study groups with regard to Ln-5gamma2-chain and MMP-13 expressions (P >0.05). CONCLUSIONS: Selective cyclooxygenase-2 inhibitor, prophylactic omega-3 fatty acid, and a combination of these two agents can inhibit gingival tissue MMP-8 expression. Moreover, the individual administration of therapeutic omega-3 may increase gingival TIMP-1 expression in contrast to no effect on MMP-8, -13, and -14 expressions in experimental periodontitis. These experimental findings in a rat model of LPS-induced periodontitis need to be verified by clinical human studies.

Wound dressing components degrade proteins detrimental to wound healing.

Excessive levels of matrix metalloproteinases (MMPs) are present in chronic wounds preventing wound closure. Reducing detrimental components may be key in healing chronic wounds. Elta Protease-containing wound dressings have been observed clinically to resolve inflammation and appear to aid healing in acute and chronic recalcitrant wounds. To investigate possible mechanisms of action, in vitro tests, zymography, collagenase assays and enzyme-linked immunosorbent assays (ELISAs), were performed to evaluate the effect of the dressing proteases on detrimental and beneficial wound healing components such as MMPs, Tissue Inhibitor of Matrix Metalloproteinases (TIMPs), cytokines and growth factors. Standards of pro- and active MMP-2, MMP-9 and chronic wound fluid (CWF) were prepared. Degradation of target proteins was enhanced by increased Elta Protease concentration, temperature and incubation time. Incubation with serial dilutions of the Elta Proteases resulted in nearly complete degradation of all MMP standards. After a 30-minute incubation, twofold diluted Elta Proteases degraded >90% of the gelatinases in CWF. ELISAs showed that Elta Proteases effectively degraded MMP-9 and tumour necrosis factor (TNF-alpha). In contrast, Platelet Derived Growth Factor (PDGF) and interleukin 1 beta were resistant to degradation by Elta Proteases. These results suggest that Elta Protease dressings appear to deactivate detrimental components in CWF, which may reduce wound bed contact with harmful proteins.

Cardioprotective role of sodium thiosulfate on chronic heart failure by modulating endogenous H2S generation.

BACKGROUND/AIMS: Sodium thiosulfate (STS) has been shown to be an antioxidant and calcium solubilizer, but the possible role of STS in dysfunctional ventricles remains unknown. Here, we assessed the effects of STS in the failing heart. METHODS: Heart failure was created by an arteriovenous fistula (AVF). Mice were divided into 4 groups: sham, AVF, sham + STS, and AVF + STS. STS (3 mg/ml) was supplemented with drinking water for 6 weeks in the appropriate surgery groups after surgery. RESULTS: M-mode echocardiograms showed ventricular contractile dysfunction with reduced aortic blood flow in AVF mice, whereas STS treatment prevented the decline in cardiac function. Ventricular collagen, MMP-2 and -9, and TIMP-1 were robustly increased with a decreasing trend in adenylate cyclase VI expression; however, STS supplementation reversed these effects in AVF mice. Among 2 enzymes that produce endogenous hydrogen sulfide (H(2)S), cystathionine-gamma-lyase (CSE) expression was attenuated in AVF mice with no changes in cystathionine-beta-synthase (CBS) expression. In addition, reduced production of H(2)S in AVF ventricular tissue was normalized with STS supplementation. Moreover, cardiac tissues were more responsive to H(2)S when AVF mice were supplemented with STS compared to AVF alone. CONCLUSIONS: These results suggested that STS modulated cardiac dysfunction and the extracellular matrix, in part, by increasing ventricular H(2)S generation.

Flavonoid glycosides isolated from Salicornia herbacea inhibit matrix metalloproteinase in HT1080 cells.

Flavonoid glycosides, isorhamnetin 3-capital O, Cyrillic-beta-d-glucoside, and quercetin 3-O-beta-d-glucoside were isolated from Salicornia herbacea and their inhibitory effects on matrix metalloproteinase-9 and -2 (MMP-9 and -2) were evaluated in human fibrosarcoma cell line (HT1080). In zymography experiments, these flavonoid glycosides led to the reduction of the expression levels and activities of MMP-9 and -2 without any significant difference between these flavonoid glycosides. Protein expression levels of both MMP-9 and MMP-2 were inhibited and TIMP-1 (tissue inhibitor of metalloproteinase-1) protein level was enhanced by these flavonoid glycosides. Moreover, a transfection study carried out with AP-1 reporter construct revealed that the reporter activity was suppressed by treatment with isorhamnetin 3-capital O, Cyrillic-beta-d-glucoside. Therefore, these results suggested that these flavonoid glycosides have a potential as valuable natural chemopreventive agents for cancer.

Amla (Emblica officinalis Gaertn.) extract promotes procollagen production and inhibits matrix metalloproteinase-1 in human skin fibroblasts.

AIM OF THE STUDY: Emblica officinalis Gaertn., commonly known as amla, is a rich dietary source of vitamin C, minerals and amino acids, and also contains various phenolic compounds. Amla extract is also known to exhibits potent antioxidant properties and to provide protection for human dermal fibroblasts against oxidative stress, and therefore it is thought to be useful for natural skin care. In this study, we investigated the effects of amla extract on human skin fibroblasts, especially for production of procollagen and matrix metalloproteinases (MMPs), in vitro. MATERIALS AND METHODS: Mitochondrial activity of human skin fibroblasts were measured by WST-8 assay. Quantification of procollagen, MMPs, and Tissue inhibitor of metalloproteinase-1 (TIMP-1) released from human skin fibroblasts were performed by immunoassay technique. RESULTS AND CONCLUSIONS: Amla extract stimulated proliferation of fibroblasts in a concentration-dependent manner, and also induced production of procollagen in a concentration- and time-dependent manner. Conversely, MMP-1 production from fibroblasts was dramatically decreased, but there was no evident effect on MMP-2. TIMP-1 was significantly increased by amla extract. From these results, it appears that amla extract works effectively in mitigative, therapeutic and cosmetic applications through control of collagen metabolism.

Effect of traditional Chinese medicine Shu-Mai-Tang on attenuating TNFalpha-induced myocardial fibrosis in myocardial ischemia rats.

Shu-Mai-Tang (SMT) is a traditional Chinese medicine for treatment of ischemic heart disease. The effect of SMT on inflammation-induced myocardial fibrosis, left ventricular (LV) remodeling, and the potential mechanism in myocardial ischemia (MI) rats were investigated. Rats with ligated left anterior descending coronary artery (MI model) were randomly divided into three groups (SMTL, SMTH, and MIR). A group undergoing Sham operation (Sham; n=16) was also included. SMT (342 or 1710 mg/kg for SMTL or SMTH groups, respectively) was orally administered daily for 1 and 6 weeks. Cardiac function, myocardial fibrosis, serum tumor necrosis factor-alpha (TNFalpha) concentration, the cardiac expressions of phosphorylated p38 MAPK and tissue inhibitor of matrix metalloproteinase (TIMP)-1 and TNFalpha were examined by echocardiography, histological staining, radioimmunoassay, western blot, respectively. In the present study, significant reduced myocardial fibrosis, as well as decreased phospho-p38 MAPK, TIMP-1, and TNFalpha proteins, and serum TNFalpha level, accompanied by improved cardiac function in the SMT-treated rats in a dose-dependent manner as compared with the MIR. These results suggested that SMT could anti-inflammation-induced myocardial fibrosis and reverse LV remodeling in MI rats, and the mechanism may be related to the effect of SMT on inhibiting p38 MAPK signaling pathway.

The effect of water extracts of Euphorbia hirta on cartilage degeneration in arthritic rats.

The effect of water extracts of Euphorbia hirta on the histological features and expressions of matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) in the rat articular cartilage was investigated. Arthritis was induced in rats using Freund's Complete Adjuvant containing heat-killed M. tuberculosis, and treated with water extracts of E. hirta. Paraffin tissue sections of the arthritic joints were evaluated. The extent of cartilage degeneration was found to be greatest in rats treated with the highest dosage of E. hirta, followed by rats in the untreated group. Rats treated with the intermediary and low dosages of Euphorbia hirta showed improved histology. MMP-13 levels were found to be decreased with decreasing dosages of E. hirta. TIMP-1 levels were found to increase with decreasing dosages of E. hirta. MMP-3 levels fluctuated without any appreciable pattern. Low dosages of E. hirta seem to be beneficial in reducing cartilage degeneration in cases of arthritis.

Piascledine modulates the production of VEGF and TIMP-1 and reduces the invasiveness of rheumatoid arthritis synoviocytes.

BACKGROUND: In rheumatoid arthritis (RA), hypertrophy of the synovial membrane generates a tumour-like pannus that invades the joint cavity and erodes cartilage and bone. Invasion of the extracellular matrix (ECM) is accompanied by angiogenesis, in which vascular endothelial growth factor (VEGF) and tissue inhibitors of metalloproteinases (TIMPs), produced by synoviocytes lining the pannus, have a primary role. Piascledine (PSD) is used in the treatment of osteoarthritis and has anti-inflammatory effects in vitro. OBJECTIVE: To study the effects of PSD on levels of VEGF and TIMP-1 and chemoinvasion in RA synoviocytes and healthy controls. METHODS: The effects of PSD 5, 10, and 20 microg/mL were evaluated, with/without interleukin-1beta (IL-1beta) and tumour necrosis factor-alpha (TNFalpha) 20 ng/mL, on synoviocytes. The levels of VEGF and TIMP-1 were assayed in the culture medium by enzyme-linked immunosorbent assay (ELISA). Chemoinvasion was measured by the Boyden chamber invasion assay. RESULTS: RA synoviocytes treated with PSD showed, compared to basal, lower levels of VEGF (41080+/-830 vs. 79210+/-920 pg/106 cells, p<0.001) and increased levels of TIMP-1 (23540+/-93.2 vs. 12860+/-42.9 ng/106 cells, p<0.001). PSD decreased dose-dependently IL-1beta and TNFalpha induced migration. CONCLUSIONS: In RA synoviocytes, and also to a lesser extent in control cells, PSD modulates VEGF and TIMP-1 and decreases chemoinvasion. PSD might have a role in the treatment of RA synovitis controlling invasiveness.

Effect of Tinospora cordifolia on the cytokine profile of angiogenesis-induced animals.

The antiangiogenic activity of Tinospora cordifolia was studied using in vivo as well as in vitro models. In vivo antiangiogenic activity was studied using B16F10 melanoma cell-induced capillary formation in animals. Intraperitoneal administration of the extract at a concentration of 20 mg/kg significantly inhibited the tumour directed capillary formation induced by melanoma cells. Analysis of the serum cytokine profile showed a drastic increase of proinflammatory cytokines such as IL-1beta, IL-6, TNF-alpha, granulocyte monocyte-colony stimulating factor (GM-CSF) and the direct endothelial cell proliferating agent vascular endothelial cell growth factor (VEGF) in the angiogenesis-induced control animals. Administration of Tinospora extract could differentially regulate these cytokine's elevation. The differential regulation is further evidenced by the increased production of antiangiogenic agents IL-2 and tissue inhibitor of metalloprotease-1 (TIMP-1) in the B16F10-injected, extract-treated animals. Moreover, using an in vitro rat aortic ring assay, it was observed that the extract at nontoxic concentrations inhibited the production of proangiogenic factors from B16F10 melanoma cells. Direct treatment of the extract also inhibits the microvessel outgrowth from the aortic ring. Hence, the observed antiangiogenic activity of the plant T. cordifolia is related, at least in part, to the regulation of the levels of these cytokines and growth factors in the blood of the angiogenesis-induced animal.