Eun-Kyung Ahn

COX-2 expression and inflammatory effects by diesel exhaust particles in vitro and in vivo.

Recent studies have shown that diesel exhaust particles (DEP) have adverse effects on the respiratory tract in vitro and in vivo, related to various pro-inflammatory cytokines and inflammatory mediators. The inflammation induced by the production of cyclooxygenase (COX)-2, an important mediator of inflammation and tumor promotion, and excess eicosanoids may be central to the pathogenesis of DEP-induced airway inflammation. However, the role of COX-2 in the pathogenesis of DEP-induced lung inflammation remains unclear, especially in vivo. In this study, we demonstrated that treatment with 50 microg/ml of DEP for 24h induced the expression of the COX-2 gene at both the transcriptional and protein levels, which led to an increase in the release of prostaglandin E(2) (PGE(2)) in A549 cells. In addition, the increased levels of COX-2 and PGE(2) by DEP exposure were significantly suppressed by treatment with 50 pg/ml of dexamethasone (Dex). We also showed that exposure to 25 mg/kg of DEP induced the expression of the COX-2 protein in mouse lung tissues, and this increased COX-2 expression was attenuated by pretreatment with 5 mg/kg of Dex. These findings suggest that COX-2 may play an important role in the pathogenesis of DEP-induced pulmonary inflammation, which is effectively inhibited by glucocorticoid treatment.

Diesel exhaust particles increase IL-1β-induced human β-defensin expression via NF-κB-mediated pathway in human lung epithelial cells

BackgroundHuman β-defensin (hBD)-2, antimicrobial peptide primarily induced in epithelial cells, is a key factor in the innate immune response of the respiratory tract. Several studies showed increased defensin levels in both inflammatory lung diseases, such as cystic fibrosis, diffuse panbronchiolitis, idiopathic pulmonary fibrosis and acute respiratory distress syndrome, and infectious diseases. Recently, epidemiologic studies have demonstrated acute and serious adverse effects of particulate air pollution on respiratory health, especially in people with pre-existing inflammatory lung disease. To elucidate the effect of diesel exhaust particles (DEP) on pulmonary innate immune response, we investigated the hBD-2 and interleukin-8 (IL-8) expression to DEP exposure in interleukin-1 beta (IL-1β)-stimulated A549 cells.ResultsIL-1β markedly up-regulated the hBD-2 promoter activity, and the subsequent DEP exposure increased dose-dependently the expression of hBD-2 and inflammatory cytokine IL-8 at the transcriptional level. In addition, DEP further induced the NF-κB activation in IL-1β-stimulated A549 cells more rapidly than in unstimulated control cells, which was showed by nuclear translocation of p65 NF-κB and degradation of IκB-α. The experiment using two NF-κB inhibitors, PDTC and MG132, confirmed that this increase of hBD-2 expression following DEP exposure was regulated through NF-κB-mediated pathway.ConclusionThese results demonstrated that DEP exposure increases the expression of antimicrobial peptide and inflammatory cytokine at the transcriptional level in IL-1β-primed A549 epithelial cells and suggested that the increase is mediated at least partially through NF-κB activation. Therefore, DEP exposure may contribute to enhance the airway-responsiveness especially on the patients suffering from chronic respiratory disease.

Diesel exhaust particles induce apoptosis via p53 and Mdm2 in J774A.1 macrophage cell line.

Diesel exhaust particles (DEP) are known to cause cardiopulmonary diseases due to their proinflammatory and cytotoxic effects. Continuous exposure to DEP potentiates chronic inflammatory processes and acute symptomatic responses in the respiratory tract. Recent studies have emphasized that alveolar cell apoptosis is a crucial step in chronic inflammation and lung injury. The phenomenon of apoptosis is a key event that successfully clears damaged cells, and its failure leads to the development of more serious diseases, such as lung cancer. The mechanism and molecular target of DEP-induced apoptosis in the respiratory tract remain unclear. In this study, J774A.1 macrophage cells were used to investigate the p53-mediated apoptotic pathway induced by DEP exposure. The results showed that murine double minute 2 (Mdm2), a negative regulator of p53, was downregulated at the protein level by DEP exposure. In contrast, the pro-apoptotic protein Bcl-2-associated X protein (Bax), an endogenous target of p53-dependent transcriptional activation, was continuously upregulated at the mRNA and protein levels by DEP exposure. Furthermore, pifithrin-alpha (p53 inhibitor) blocked DEP-induced apoptosis as well as p53 activation. Taken together, the findings of the present study suggest that DEP trigger apoptosis in J774A.1 macrophage cells via the activation of p53, followed by Bax.

Broussonetia kazinoki modulates the expression of VEGFR-2 and MMP-2 through the inhibition of ERK, Akt and p70S6K‑dependent signaling pathways: Its implication in endothelial cell proliferation, migration and tubular formation

Broussonetia kazinoki (BK) has been used as a traditional medicine to improve vision, as well as for inflammatory and infectious diseases. In the present study, we investigated the effects and molecular mechanism of the ethanolic extract of BK on cell proliferation, migration and tubular formation in vascular endothelial growth factor-A (VEGF-A)-treated human umbilical vein endothelial cells. BK treatment inhibited VEGF-A-stimulated endothelial cell proliferation through the downregulation of cell cycle-related proteins including cyclin-dependent kinases and cyclins. Moreover, BK treatment suppressed cell migration and tubular formation in response to VEGF-A. These anti-angiogenic activities of BK were associated with the inactivation of mitogenic signaling pathways including extracellular signal-regulated kinase, Akt and p70S6K, and the subsequent downregulation of VEGFR-2 and matrix metalloproteinase-2. Taken together, these findings suggest further evaluation and development of BK as a potential therapeutic agent for the treatment and prevention of angiogenesis-related diseases including cancer.

A mechanistic study on the anti-cancer activity of ethyl caffeate in human ovarian cancer SKOV-3 cells

In the present study, we investigated the effect and molecular mechanism of ethyl caffeate (EC), a natural phenolic compound isolated from Ligularia fischeri, on human ovarian cancer cell proliferation and progression. EC-mediated inhibition of cell proliferation in SKOV-3 cells was accompanied by reduced expression of cell cycle-related proteins such as cyclin-dependent kinases and cyclins, resulting in pRb hypophosphorylation and G₁ phase cell cycle arrest. Moreover, EC treatment markedly inhibited cell migration and invasion. These regulatory effects of EC on ovarian cancer cell proliferation, migration and invasion were associated with inactivation of mitogenic signaling pathways such as Akt, ERK and p38(MAPK), and down-regulation of cell surface signaling molecules including receptor tyrosine kinases, integrin α3β1 and N-cadherin. Taken together, these findings suggest further evaluation and development of EC for the treatment and prevention of ovarian cancer.

Marmesin is a novel angiogenesis inhibitor: Regulatory effect and molecular mechanism on endothelial cell fate and angiogenesis

In the present study, we investigated the effects and molecular mechanism of marmesin, a coumarin compound isolated from Broussonetia kazinoki, on vascular endothelial growth factor-A (VEGF-A)-induced endothelial cell responses in vitro and angiogenic sprouting in aortic rings ex vivo. Marmesin treatment inhibited VEGF-A-stimulated endothelial cell proliferation through down-regulation of cell cycle-related proteins including cyclin-dependent kinases and cyclins, leading to pRb hypophosphorylation and G1 phase cell cycle arrest. In addition, marmesin treatment abrogated VEGF-A-induced endothelial cell migration, invasion and capillary-like structure formation in vitro as well as angiogenic sprouting ex vivo. These anti-angiogenic activities of marmesin were mediated through inactivation of VEGF-A-stimulated signaling pathways, and down-regulation of cell surface signaling molecules including VEGF receptor-2, human epidermal growth factor receptor-2, integrin β1 and integrin-liked kinase. Taken together, these findings clearly support the pharmacological roles of marmesin in regulating angiogenesis, and warrant further evaluation and development as a potential therapeutic agent for the treatment and prevention of angiogenesis-related diseases including cancer.

Neolignan inhibitors of antigen-induced degranulation in RBL-2H3 cells from the needles of Pinus thunbergii

The ethanolic extract of the needles of Pinus thunbergii was found to suppress antigen mediated degranulation of rat basophilic leukemia (RBL-2H3) cells. A new neolignan glycoside, named pinusthunbergiside A (1), as well as six known neolignan glycosides (2-7) were isolated from the ethanolic extract using bioassay-guided fractionation. Their structures were elucidated by a combination of 1D and 2D NMR, HRESI-MS, and circular dichroism (CD) data. Compounds 2-7 were found for the first time in this plant. The inhibitory effects of isolated constituents on the release of β-hexosaminidase from RBL-2H3 cells were examined, and compounds 2, 3, 5, and 6 were found to show the inhibitory activity with IC₅₀ values ranging between 52.3 and 75.3 μM.

2,8-Decadiene-1,10-Diol Inhibits Lipopolysaccharide-Induced Inflammatory Responses Through Inactivation of Mitogen-Activated Protein Kinase and Nuclear Factor-κB Signaling Pathway.

Amomum tsao-ko (A. tsao-ko) has been used as a traditional medicine for the treatment of infectious and digestive disorders. In the present study, we report the anti-inflammatory activity and molecular mechanism of 2,8-decadiene-1,10-diol (DDO) isolated from the extract of A. tsao-ko in lipopolysaccharide-stimulated RAW 264.7 cells. DDO treatment inhibited the production of nitric oxide and prostaglandin E2 by downregulating inducible nitric oxide synthase and cyclooxygenase-2 expression, respectively. Moreover, DDO suppressed the production of pro-inflammatory cytokines such as interleukin-6 and tumor necrosis factor-α. These inhibitory effects of DDO on the expression of inflammatory proteins were found to be mediated through the inactivation of mitogen-activated protein kinases (MAPKs) such as extracellular signal-regulated kinase, c-Jun-N-terminal kinase and p38MAPK, and inhibition of nuclear factor-κB (NF-κB) pathways including degradation of inhibitor of κB-α and nuclear localization of NF-κB. Taken together, these findings demonstrate the pharmacological roles and molecular mechanisms of DDO in regulating inflammatory responses, and suggest further evaluation and development of DDO as a potent therapeutic agent for the treatment of inflammatory disorders.

Ligularia fischeri inhibits endothelial cell proliferation, invasion and tube formation through the inactivation of mitogenic signaling pathways and regulation of vascular endothelial cadherin distribution and matrix metalloproteinase expression

Ligularia fischeri (LF) has been used as an edible herb and traditional medicine for the treatment of inflammatory and infectious diseases. In the present study, we report the effects and molecular mechanism of the ethanolic extract of LF on cell proliferation, invasion and tube formation in human umbilical vein endothelial cells (HUVECs). LF-mediated inhibition of cell proliferation was accompanied by reduced expression of cell cycle-related proteins such as cyclin-dependent kinases (Cdks) and cyclins, leading to pRb hypophosphorylation and G1 phase cell cycle arrest. We also show that LF treatment inhibited cell invasion and tube formation in HUVECs. These anti-angiogenic activities of LF were associated with the inactivation of mitogenic signaling pathways, induction of vascular endothelial (VE)-cadherin distribution at cell-cell contacts and inhibition of matrix metalloproteinase (MMP) expression. Collectively, our findings demonstrate the pharmacological functions and molecular mechanisms of LF in regulating endothelial cell fates, and support further development as a potential therapeutic agent for the treatment and prevention of angiogenesis-related disorders including cancer.

Anti-obesity effects of Clausena excavata in high-fat diet-induced obese mice

Clausena excavata (C. excavata) has been used as a traditional medicine for the treatment of abdominal pain, enteritis, dysentery, and malaria. The present study was designed to evaluate the effect of a 50% ethanol extract of C. excavata (ECE) on weight loss, adipocyte size, and obesity-related biochemical parameters in high-fat diet (HFD)-induced obese mice. After 6 weeks of HFD + ECE administration, HFD-induced total fat, subcutaneous fat, and visceral fat were evaluated by micro-computed tomography. The serum levels of triglyceride (TG), total cholesterol (TCH), high-density lipoprotein (HDL) cholesterol, and low-density lipoprotein (LDL) cholesterol were evaluated with a biochemical analyzer, and leptin and adiponectin levels in the serum were assessed via enzyme-linked immunoassay (ELISA). Moreover, adipocyte size and lipid formation in the liver were examined. We found that weight gain, epididymal fat pad weight, adipocyte size, and lipid formation were markedly attenuated in the livers of HFD-induced obese mice treated with ECE. Furthermore, TG, TCH, and leptin decreased in the serum, whereas adiponectin increased. In conclusion, our data show that ECE has potent anti-obesity activity in vivo and support the development of ECE as a potential therapeutic agent for the treatment of obesity.