Anti-senescence and anti-wrinkle activities of 3-bromo-4,5-dihydroxybenzaldehyde from Polysiphonia morrowii Harvey in human dermal fibroblasts

Objective:To investigate the anti-senescence effect of 3-bromo-4,5-dihydroxybenzaldehyde (BDB) from Polysiphonia morrowii Harvey in human dermal fibroblasts (HDF). Methods:HDF were subjected to treatment of BDB and then treated with hydrogen peroxide (H2O2) to induce premature senescence. Senescence-associated β-galactosidase (SA-β-gal) activity in HDF was determined using the SA-β-gal staining method. Intracellular reactive oxygen species (ROS) production was measured using the 2',7'-dichlorodihydrofluorescein diacetate assay. Western blotting assay was performed to assess the level of antioxidant enzyme glutathione peroxidase 1 (GPX1). In addition, intracellular collagen and collagenase contents were analyzed using the respective ELISA kits. Elastase activity in HDF supernatants was measured from p-nitroaniline release and normalized using total protein content. Results:Treatment of HDF with H2O2 increased the activity of SA-β-gal, but BDB pre-treatment resulted in the reduction of SA-β-gal activity. Moreover, BDB significantly reduced H2O2-induced intracellular ROS production. BDB also markedly increased the level of GPX1, which was inhibited by 400 μM of H2O2. Furthermore, in in vitro study, BDB significantly increased intracellular collagen content and decreased matrix metalloproteinase-1 and elastase activities in HDF. Conclusions:Our results demonstrate that BDB shows anti-senescence and anti-wrinkle activities in vitro.

Anti-aging effects of Muntingia calabura leaves extract in D-galactose-induced skin aging mouse model

Glycation and production of free radicals become important mechanisms underlying skin aging. Muntingia calaburais reported to have antioxidant activity in many studies. The effects of M. calabura aqueous leaves extract (MCALE)on oxidative stress and histological changes of mouse model of skin aging were evaluated in this research. Twentymale albino mice were divided into five groups: healthy control; aging control; aging + MCALE 35 mg/kg; aging +MCALE 70 mg/kg; and aging+vitamin C 28 mg/kg. To induce aging condition, oral gavage of D-galactose 500 mg/kg/day were given for 6 weeks. Prior to treatment, blood samples were taken for malondealdehyde (MDA) analyses.MCALE and vitamin C were administered subsequently by oral gavage for 4 weeks and at the end, MDA analyseswere performed again. Routine and van Gieson’s staining were performed to analyze epidermal thickness, fibroblastcell, and density of dermal collagen. Groups received MCALE 70 mg/kg and vitamin C had lower plasma MDAlevel; higher fibroblast number and density of collagen bundles which is reduced in the aging group (p < 0.05).However, epidermal thickness among the five groups was not significantly different. It was concluded that MCALEhad antioxidant and anti-aging effects on D-galactose-induced mouse model of skin aging

Effects of Remifentanil Preconditioning Attenuating Oxidative Stress in Human Dermal Fibroblast

Human dermal fibroblast is essential in wound healing of the skin through the synthesis of extracellular matrix proteins. With respect to oxidative stress, the effects of remifentanil on human dermal fibroblast have received little attention. Therefore, we investigated the effects of remifentanil on the apoptosis and autophagic reaction of human dermal fibroblasts under oxidative stress. The subjects were divided into the following groups: Control group: cells were incubated at 37℃ in a humidified atmosphere with 5% CO₂. Hydrogen peroxide (H₂O₂) group: cells were exposed to H₂O₂ for 2 h. RPC/H₂O₂ group: cells were pretreated with remifentanil for 2 h and exposed H₂O₂ for 2 h. 3-MA/RPC/H₂O₂ group: cells were pretreated with 3-methyladenine (3-MA) and remifentanil for 1 h and 2 h, respectively. We measured cell viability using MTT assay. Western blot analysis was used to determine the expression levels of proteins associated with apoptosis and autophagy. Quantification of apoptotic cells was performed using flow cytometer analysis, and autophagic vacuoles were observed under a fluorescence microscope. Remifentanil treatment increased the proliferation of human dermal fibroblast and decreased apoptotic cell death, enhancing autophagic activity under oxidative stress. However, 3-MA, the autophagy pathway inhibitor, inhibited the protective effect of remifentanil in oxidative stress. This study demonstrates that remifentanil activated autophagy and decreased apoptotic death of human dermal fibroblasts under oxidative stress. Our results suggest that remifentanil may help in the treatment of oxidative stress.

Reduced Expression of YAP in Dermal Fibroblasts is Associated with Impaired Wound Healing in Type 2 Diabetic Mice

Dermal fibroblasts play essential roles in wound healing and their dysfunction has been shown to be associated with impaired wound healing in diabetes. In the present study, we aimed at investigating whether Yes-associated protein (YAP), a mediator of mechanotransduction in dermal fibroblasts, is associated with impaired wound healing in diabetic mice. Compared with that in the control, the rate of wound contraction was decreased twofold in db/db type 2 diabetic mice (db/db mice). To mimic diabetic pathological condition, dermal fibroblasts were cultured under high glucose conditions (25.5 mM glucose). Further, dermal fibroblast-mediated contraction of wound was evaluated by in vitro collagen gel contraction assay. Dermal fibroblasts cultured under hyperglycemic condition showed impaired gel contraction and mitochondrial dysfunction, compared to the cells cultured under normoglycemic conditions (5.5 mM glucose). Importantly, compared with the normal dermal fibroblasts, diabetic db/db dermal fibroblasts expressed lower levels of growth factors and cytokines that enhance wound healing, such as insulin-like growth factor-1, stromal cell-derived factor-1, connective tissue growth factor, and transforming growth factor-β (TGF-β). The quantity of YAP mRNA was also lower in diabetic db/db dermal fibroblasts, compared with that in the control fibroblasts. These results indicate that impaired wound healing in diabetics is associated with the dysfunction of dermal fibroblasts, including downregulation of YAP, which plays essential roles in extracellular matrix remodeling and TGF-β-mediated wound healing.

Dermal fibroblast expression of stromal cell-derived factor-1 (SDF-1) promotes epidermal keratinocyte proliferation in normal and diseased skin

Stromal cells provide a crucial microenvironment for overlying epithelium. Here we investigated the expression and function of a stromal cell-specific protein, stromal cell-derived factor-1 (SDF-1), in normal human skin and in the tissues of diseased skin. Immunohistology and laser capture microdissection (LCM)-coupled quantitative real-time RT-PCR revealed that SDF-1 is constitutively and predominantly expressed in dermal stromal cells in normal human skin in vivo. To our surprise, an extremely high level of SDF-1 transcription was observed in the dermis of normal human skin in vivo, evidenced by much higher mRNA expression level than type I collagen, the most abundant and highly expressed protein in human skin. SDF-1 was also upregulated in the tissues of many human skin disorders including psoriasis, basal cell carcinoma (BCC), and squamous cell carcinoma (SCC). Double immunostaining for SDF-1 and HSP47 (heat shock protein 47), a marker of fibroblasts, revealed that fibroblasts were the major source of stroma-cell-derived SDF-1 in both normal and diseased skin. Functionally, SDF-1 activates the ERK (extracellular-signal-regulated kinases) pathway and functions as a mitogen to stimulate epidermal keratinocyte proliferation. Both overexpression of SDF-1 in dermal fibroblasts and treatment with rhSDF-1 to the skin equivalent cultures significantly increased the number of keratinocyte layers and epidermal thickness. Conversely, the stimulative function of SDF-1 on keratinocyte proliferation was nearly completely eliminated by interfering with CXCR4, a specific receptor of SDF-1, or by knock-down of SDF-1 in fibroblasts. Our data reveal that extremely high levels of SDF-1 provide a crucial microenvironment for epidermal keratinocyte proliferation in both physiologic and pathologic skin conditions.

Effect of supplementation of dermal fibroblasts conditioned medium on expansion of keratinocytes through enhancing attachment.

In the present study in vitro expansion of human keratinocytes by supplementing dermal fibroblasts conditioned medium (DFCM) has been reported. Effect of two different DFCM acquired by culturing fibroblasts in keratinocyte-specific medium (defined keratinocytes serum free medium, DFCM-DKSFM) and fibroblast-specific serum free medium (F12: DMEM nutrient mix, DFCM-FD) have been compared. Growth kinetics of keratinocytes in terms of efficiency of cell attachment, expansion index, apparent specific growth rate and growth potential at the end of culture was evaluated in culture supplemented with DFCM-DKSFM and DFCM-FD in comparison with control i.e. DKSFM only. Results indicated that supplementation of DFCM caused significant increase in keratinocyte attachment. Efficiency of keratinocyte attachment in culture supplemented with DFCM-DKSFM was significantly higher compared to those cultured in DFCM-FD and DKSFM. In addition, the expansion index of keratinocytes in cultures supplemented with DFCM-DKSFM and DFCM-FD were 3.7 and 2.2 times higher than that of control condition even though the apparent growth rate and proliferative potential was found significantly lower. These results suggested that supplementation of DFCM enhanced expansion of keratinocyte by increasing efficiency of cell attachment, and DFCM-DKSFM provided suitable condition for in vitro expansion of keratinocytes compared to DFCM-FD and control condition.

Effects of Hepatocyte Growth Factor on Collagen Synthesis and Matrix Metalloproteinase Production in Keloids

Keloids are pathologic proliferations of the dermal layer of the skin resulting from excessive collagen production and deposition. Hepatocyte growth factor (HGF) increases the expression of matrix metalloproteinase (MMP)-1 and suppresses collagen synthesis to modulate extracellular matrix turnover. To investigate the anti-fibrotic effects of HGF, we examine the mRNA expression of collagen types I and III and matrix metalloproteinase (MMP-1, MMP-3) on human dermal fibroblast (HDF) cell lines and keloid fibroblasts (KFs, n = 5) after adding various amount of HGF protein. We also evaluated the enzymatic activity of MMP-2, MMP-9 by zymograghy. In HDFs treated with TGF-beta1 and HGF protein simultaneously, both type I and III collagen mRNA expression significantly decreased (P < 0.05). Expression of MMP-1, MMP-3 mRNA also decreased. However, the mRNA expression of MMP-1, MMP-3 significantly increased in KFs with increasing amount of HGF in dose dependent manner (P < 0.05). The enzymatic activities of MMP-2 increased with increasing HGF protein in a dose-dependent manner. However, the enzymatic activity of MMP-9 did not change. These results suggest that the anti-fibrotic effects of HGF may have therapeutic effects on keloids by reversing pathologic fibrosis.

Royal jelly enhances migration of human dermal fibroblasts and alters the levels of cholesterol and sphinganine in an in vitro wound healing model

Oral administration of royal jelly (RJ) promotes wound healing in diabetic mice. Concerns have arisen regarding the efficacy of RJ on the wound healing process of normal skin cells. In this study, a wound was created by scratching normal human dermal fibroblasts, one of the major cells involved in the wound healing process. The area was promptly treated with RJ at varying concentrations of 0.1, 1.0, or 5 mg/ml for up to 48 hrs and migration was analyzed by evaluating closure of the wound margins. Furthermore, altered levels of lipids, which were recently reported to participate in the wound healing process, were analyzed by HPTLC and HPLC. Migration of fibroblasts peaked at 24 hrs after wounding. RJ treatment significantly accelerated the migration of fibroblasts in a dose-dependent manner at 8 hrs. Although RJ also accelerated the migration of fibroblasts at both 20 hrs and 24 hrs after wounding, the efficacy was less potent than at 8 hrs. Among various lipid classes within fibroblasts, the level of cholesterol was significantly decreased at 8 hrs following administration of both 0.1 ug/ml and 5 mg/ml RJ. Despite a dose-dependent increase in sphinganines, the levels of sphingosines, ceramides, and glucosylceramides were not altered with any concentration of RJ. We demonstrated that RJ enhances the migration of fibroblasts and alters the levels of various lipids involved in the wound healing process.

Expression of Sodium-dependent Vitamin C Transporter in Rat Dermal Fibroblasts / 대한피부과학회지

BACKGROUND: Vitamin C is one of the most typical types of water-soluble antioxidants that exerts a variety of biochemical actions on a living body. It acts on the skin by promoting wound healing, preventing skin aging, and inhibiting skin cancer. It also works not only as an antioxidant, protecting the skin from UV radiation but also as an anti-inflammatory agent. It reinforces immunity as well. Recent studies proved the whitening effect of vitamin C, and it can be instilled into the skin by way of iontophoresis. When vitamin C is transported in vivo it is either by simple diffusion or by a transporter. Only a small amount is transported by simple diffusion and the transporter is responsible for most of the vitamin C transport. This study was designed to evaluate the presence of sodium dependent vitamin C transporter (SVCT) and to identify which factor controls its expression. METHODS: Expressions of SVCT 1 and 2 mRNA in the rats' dermal fibroblast were measured by RT-PCR at 3, 6, 12, 24, and 48 hours. RESULTS: The results were used to compare the expression levels of SVCT-1 and SVCT-2 when treated with TGF-beta, estradiol, and retinoic acid. Estradiol showed the highest level of expression of SVCT-1 and SVCT-2. The next highest was TGF-beta, followed by retinoic acid. CONCLUSION: SVCT-1 and SVCT-2 were found to be expressed in the rats' dermal fibroblasts, and exposure to estradiol, TGF-beta and retinoic acid resulted in a higher degree of their expression.

Hypoxia Induces the Expression of Connective Tissue Growth Factor in Dermal Fibroblasts / 대한류마티스학회지

OBJECTIVE: Connective tissue growth factor (CTGF) has been proposed to play a role in fibrotic process of systemic sclerosis. Since hypoxia was known to be associated with fibrosis in several profibrogenic conditions, we investigated whether CTGF expression in dermal fibroblast is regulated by hypoxia caused by microvascular loss. METHODS: Dermal fribroblasts from patient with systemic sclerosis and normal controls were cultured in the presence of cobalt chloride (CoCl2), a chemical inducer of HIF-1alpha or hypoxic culture conditions. Expression of HIF-1alpha, VEGF and CTGF was evaluated by semiquantitative reverse transcription-polymerase chain reaction and Western blotting. RESULTS: Scleroderma fibroblasts expressed increased levels of HIF-1alpha, VEGF and CTGF compared to normal dermal fibroblasts. Dermal fibroblasts exposed to various concentration of CoCl2 (1~100microM) enhanced the expression of CTGF mRNA in dose-dependent fashion. Actinomycin D significantly blocked the hypoxia-mediated up-regulation of CTGF mRNA expression, whereas cycloheximide did not block the up-regulation. Up-regulation of CTGF by hypoxia was not mediated by endogenous production of transforming growth factor (TGF)-beta. In time-kinetics study, dermal fibroblasts from scleroderma patients exhibited earlier peak expression of CTGF mRNA than those from normal dermal fibroblasts. In addition, simultaneous treatment of suboptimal concentration of CoCl2 and TGF-beta exhibited the up-regulation of CTGF mRNA in additive fashion. Interferon-gamma did not modulate the expression of CTGF mRNA induced by CoCl2, while the up-regulation of CTGF by TGF-beta was downregulated by Interferon-gamma in a dose-dependent fashion. CONCLUSION: These data indicate that hypoxia up-regulates the expression of CTGF in dermal fibroblasts and provide the evidence that hypoxia caused by microvascular alterations contributes the progression of fibrosis in systemic sclerosis by up-regulation of CTGF.

The Effects of Epigallocatechin Gallate on Ultraviolet B Irradiated Cultured Human Skin Fibroblasts / 대한피부과학회지

BACKGROUND: The main polyphenol components in green tee are (-)-epicatechin, (-)-epigallocatechin, (-)-epicatechin gallate and (-)-epigallocatechin gallate (EGCG). It is well known that flavonoids such as catechins can be protective against inflammatory and cancer and cardiovascular diseases. These protective effects are largely due to their inhibition of some enzymes and antioxidative activities by scavenging free radicals. Ultraviolet(UV) exposure of the skin, particulary UVB (290-320nm), causes adverse biological effects, including alterations in cutaneous immune cells, photoaging and photocarcinogenesis. Several studies have shown that EGCG afforded protection against UVB-induced inflammatory responses and photocarcinogenesis in murine models. OBJECTIVE AND METHODS: In this study, we investigated the effects of EGCG on UVB irradiated human skin fibroblasts using viability test, thiobarbituric acid assay, propidium iodide(PI) stain, and western blot analyses and RT-PCR. RESULTS: Cell survival curves after UVB irradiation showed dose dependent decrement pattern by trypan blue exclusion assay. Only 42% of dermal fibroblasts survived at 150 mJ/cm2 UVB irradiation. The damage was associated with cell membrane lipid peroxidation, as shown by accumulation malondialdehyde(MDA). By pre-cultivation with EGCG (50nmol), a significant preventive effect was noted on the increase in the absolute number of surviving cells(up to 81.5% of cells survived) and the levels of MDA markedly decreased. Morphological changes associated with apoptotic cell death were easily distinguished by PI stain. Bases on our finding, we investgated the regulation of p53, p21, bax, bcl-2, cyclin D1, E, Cdk2, and PARP proteins by western blot analyses. The expression p53 protein was elevated by following UVB exposure which was inhibited by EGCG treatment. Using RT-PCR, the transcription of p53, fas and jun gene showed similar results which obtained by western blot analyses. CONCLUSION: EGCG, which have newly accepted as a potential UV protection properties, is effective membrane peroxidation inhibitor and prevent apoptotic changes when present in relevant concentration at the site of action beginning and during UVB irradiation. And the protective mechanism of EGCG against UVB-induced cell damage maybe, at least in part, related with p53, fas and jun pathway.

The Effect of Granulocyte-Macrophage Colony-Stimulating Factor on Human Dermal Fibroblast Proliferation

Granulocyte-Macrophage colony-stimulating factor(GM- CSF) is naturally generated protein that stimulates the survival, proliferation, and differentiation of myeloid progenitor cells. The determination of the molecular sequence of this protein by recombinant DNA technology enabled us to produce sufficient quantity for preclinical and clinical use. In the animal studies, rhGM-CSF to wounds has been reported to result in increased formation of granulation tissue, increased breaking strength, and reversal of wound contraction. A number of case reports have been published on the favorable effect of rhGM-CSF as a treatment for infected, nonhealing wounds, and ulcers. However, there are no clinical reports about the effect of GM-CSF on wound healing in normal patients. Therefore, in this report, we examined the effect of GM-CSF on the proliferation of human dermal fibroblasts which play a crucial role in wound healing process in vitro. To determine an optimal GM-CSF concentration for human dermal fibroblast proliferation, the cells were incubated with either one of 13 concentrations of GM-CSF(0 - 30mug/ml). The media used in this study was DMEM/F- 12(GIBCO, Grand Island, NY, USA). The fibroblasts were seeded at 1.5 x 104 cells/well in 500mul of medium including 10% fetal bovine serum and either one of 13 concentrations of GM-CSF in 24-well plates. The cells were incubated for 6 days at 5% CO2, 100% humidity at 37degrees C. On the 6th day of plating, fibroblast proliferation was determined by hematocytometer. Each concentration was tested 8 times.Low concentration of GM-CSF(below 5.0mug/ml) stimulated the proliferation of human dermal fibroblasts. How ever, high concentration of GM-CSF(over 10mug/ml) downregulated the proliferation of human dermal fibroblasts. The best fibroblast proliferation was seen at 1.0mug/ml of GM- CSF. These results demonstrated that GM-CSF influenced human dermal fibroblast proliferation and the GM-CSF concentration was critically important factor in vitro.

The Effect of Granulocyte-Macrophage Colony-Stimulating Factor on Human Dermal Fibroblast Proliferation

Granulocyte-Macrophage colony-stimulating factor(GM- CSF) is naturally generated protein that stimulates the survival, proliferation, and differentiation of myeloid progenitor cells. The determination of the molecular sequence of this protein by recombinant DNA technology enabled us to produce sufficient quantity for preclinical and clinical use. In the animal studies, rhGM-CSF to wounds has been reported to result in increased formation of granulation tissue, increased breaking strength, and reversal of wound contraction. A number of case reports have been published on the favorable effect of rhGM-CSF as a treatment for infected, nonhealing wounds, and ulcers. However, there are no clinical reports about the effect of GM-CSF on wound healing in normal patients. Therefore, in this report, we examined the effect of GM-CSF on the proliferation of human dermal fibroblasts which play a crucial role in wound healing process in vitro. To determine an optimal GM-CSF concentration for human dermal fibroblast proliferation, the cells were incubated with either one of 13 concentrations of GM-CSF(0 - 30mug/ml). The media used in this study was DMEM/F- 12(GIBCO, Grand Island, NY, USA). The fibroblasts were seeded at 1.5 x 104 cells/well in 500mul of medium including 10% fetal bovine serum and either one of 13 concentrations of GM-CSF in 24-well plates. The cells were incubated for 6 days at 5% CO2, 100% humidity at 37degrees C. On the 6th day of plating, fibroblast proliferation was determined by hematocytometer. Each concentration was tested 8 times.Low concentration of GM-CSF(below 5.0mug/ml) stimulated the proliferation of human dermal fibroblasts. How ever, high concentration of GM-CSF(over 10mug/ml) downregulated the proliferation of human dermal fibroblasts. The best fibroblast proliferation was seen at 1.0mug/ml of GM- CSF. These results demonstrated that GM-CSF influenced human dermal fibroblast proliferation and the GM-CSF concentration was critically important factor in vitro.

Chronic Hypotony Model in the Rabbit

Chronic hypotony is an importnat cause of functional failure after proliferative vitreoretinopathy(PVR) surgery even if the retina is successfully reattached. The purpose of this study was to create a relevant model of chronic hypotony in the rabbit. Eighteen pigmented rabbits weighing 3~4kg were used in this experiment. Pars plana lensectomy was performed on fourteen eyes of fourteen rabbits with a fragmatome and a vitreous cutter. At the end of surgery, 0.2ml of phosphate-buffered saline(PBS) containing 100,000 cells of cultured rabbit dermal fibroblasts was injected over the epiciliary area. As a control, pars plana lensectomy alone was done on four eyes of four rabbits. The intraocular pressure(IOP) was measured on days 7, 14, 21, 28. All fourteen eyes following lensectomy and fibroblast injection had an IOP less than 5mmHg on each follow-up and a mean of 2.5+/-0.6mmHg (mean+/-S.D.) on day 28. Four control eyes with lensectomy alone had an IOP of 7.5+/-2.1mmHg(mean+/-S.D.) on day 28 (P<0.05). On gross examination, the development of fibrous translucent epiciliary membrane was found on day 28. Microscopic examination of eyes obtained on day 28 showed changes in the ciliary epithelium that included an absence or atrophy of the non-pigmented ciliary epithelium, an atrophy and cystic vacuolization of the pigmented ciliary epitheliu, and the interstitial edema of the ciliary body stroma. In conclusion, a model of chronic hypotony with epiciliary membrane using cultured fibroblasts was created in the rabbit. This model may be useful to help elucidate the pathophysiology of chronic hypotony and to investigate potential treatments.

The Effect of Daunorubicin on Experimental Proliferative Vitreoretinopathy

Proliferative vitreoretinopathy (PVR) is a main cause of failure in retinal reattachment surgery. There have been many studies about the inhibition of proliferative vitreoretinophthy with several drugs. Authors investigated the inhibitory effect of proliferative vitreoretinopathy and retinal toxicity with various concentration of daunorubicin after intravitreal injection into the eyes of the pigmented rabbit. 7 pigment rabbit (11eyes) were used as subjects. After lensectomy and vitrectomy, control group was injected dermal fibroblast and F-BSS, and treatment group was injected dermal fibroblast and 5, 10, 15, 30 nmol Daunorubicin. At two weeks after intravitreal injection, both group were enucleated and examined with gross finding, light--microscopy, and electronmicroscopy. In all control group, proliferative vitreoretinopathy was found, but only preretinal membrane formation was found in 5, 10 nmol Daunorubicin injected group. In 15 nmol Daunorubicin injected group, the retina structure was preserved normally. In 30 nmol Daunorubicin injected group, the retinal outer segment was degenerated in microscopic finding. These results show that Daunorubicin has a potent effect on proliferative vitreoretinopathy, especially in 15 nmol, but retinal toxicity is suspected in marethan 30 nmol.

Effect of paeonol on the expression of MMP-9 mRNA and cytokines production in human skin fibroblasts induced by TNF-? / 中国药理学通报

Aim To explore the possible anti-inflammation mechanism of paeonol by investigating its effects on the MMP-9 mRNA expression and cytokines production in human dermal fibroblasts induced by TNF-?.Methods The effect of paneonol on the expression of MMP-9 mRNA was detected by semi-quantitative RT-PCR.The modulation of paneonol on the production of IL-1?,IL-6 and IL-8 in fibroblasts was measured by ELISA.Results MMP-9 hardly expressed in human dermal fibroblast.The results also showed that TNF-? significantly induced the expression of MMP-9 in fibroblasts and at the same time paeonol inhibited the expression of MMP-9.TNF-? stimulated the production of IL-1? and IL-8 in fibroblasts,while 10~100 mg?L-1 paeonol inhibited TNF-?-induced IL-1? and IL-8 production in fibroblasts but had nothing to do with the production of IL-6.Conclusions Paeonol can inhibit the expression of MMP-9 mRNA,IL-1? and IL-8 induced by TNF-?.