Project description:BACKGROUND:Oral lichen planus (OLP) is known as a chronic inflammatory disease. Our recent studies have suggested that vitamin D/vitamin D receptor (VDR) signaling exerts its protective effects on oral keratinocyte apoptosis by regulating microRNA-802 and p53-upregulated modulator of apoptosis (PUMA), but its roles in oral epithelial inflammatory responses in OLP are still unknown. Herein, we identify lipopolysaccharide (LPS) is able to enhance interferon gamma (IFNγ) and interleukin-1 beta (IL-1β) productions in human oral keratinocytes (HOKs) dependent on hypoxia-inducible factor-1α (HIF-1α). METHODS:HIF-1α and cytokines levels in HOKs were investigated by real-time PCR and western blotting after LPS challenge. The effects of 1,25(OH)2D3 on LPS-induced HIF-1α and cytokines were tested by real-time PCR, western blotting, siRNA-interference and plasmids transfection techniques. The roles of 1,25(OH)2D3 in regulating HIF-1α levels were investigated using western blotting, siRNA-interference, plasmids transfection and Chromatin Immunoprecipitation (ChIP) assays. Finally, HIF-1α, IFNγ and IL-1β expressions in oral epithelia derived from mice and individuals were measured by real-time PCR, western blotting and immunohistochemical staining. RESULTS:As a critical regulator, vitamin D suppresses LPS-induced HIF-1α to block IFNγ and IL-1β productions. Mechanistically, vitamin D inactivates nuclear factor-κB (NF-κB) pathway and up-regulates von Hippel-Lindau (VHL) levels, leading to HIF-1α reduction. Moreover, HIF-1α status of oral epithelia is elevated in VDR-/- mie as well as in VDR-deficient human biopsies, accompanied with increased IFNγ and IL-1β. CONCLUSION:Collectively, this study uncovers an unrecognized roles of vitamin D/VDR signaling in regulating cytokines in oral keratinocytes and reveals the molecular basis of it.
Project description:Catalpa ovata (Bignoniaceae) is widely distributed throughout Korea, China, and Japan. This study investigated the anti-inflammatory effects of catalpalactone isolated from C. ovata in lipopolysaccharide (LPS)-induced RAW264.7 cells. Catalpalactone significantly inhibited nitric oxide (NO) production and inducible NO synthase (iNOS) expression in LPS-induced RAW264.7 cells. The levels of cytokines such as interleukin-6 and tumor necrosis factor-α were reduced under catalpalactone exposure in LPS-induced RAW264.7 cells. Additionally, catalpalactone suppressed signal transducer and activator of transcription 1 (STAT-1) protein expression and interferon-β (IFN-β) production. Treatment with catalpalactone prevented interferon regulatory factor 3 (IRF3) and nuclear factor-κB (NF-κB) activation. Taken together, these results suggest that the anti-inflammatory effects of catalpalactone are associated with the suppression of NO production and iNOS expression through the inhibition of IRF3, NF-κB, and IFN-β/STAT-1 activation.
Project description:Punicalagin, a hydrolysable tannin of pomegranate juice, exhibits multiple biological effects, including inhibiting production of pro-inflammatory cytokines in macrophages. Autophagy, an intracellular self-digestion process, has been recently shown to regulate inflammatory responses. In this study, we investigated the anti-inflammatory potential of punicalagin in lipopolysaccharide (LPS) induced RAW264.7 macrophages and uncovered the underlying mechanisms. Punicalagin significantly attenuated, in a concentration-dependent manner, LPS-induced release of NO and decreased pro-inflammatory cytokines TNF-? and IL-6 release at the highest concentration. We found that punicalagin inhibited NF-?B and MAPK activation in LPS-induced RAW264.7 macrophages. Western blot analysis revealed that punicalagin pre-treatment enhanced LC3II, p62 expression, and decreased Beclin1 expression in LPS-induced macrophages. MDC assays were used to determine the autophagic process and the results worked in concert with Western blot analysis. In addition, our observations indicated that LPS-induced releases of NO, TNF-?, and IL-6 were attenuated by treatment with autophagy inhibitor chloroquine, suggesting that autophagy inhibition participated in anti-inflammatory effect. We also found that punicalagin downregulated FoxO3a expression, resulting in autophagy inhibition. Overall these results suggested that punicalagin played an important role in the attenuation of LPS-induced inflammatory responses in RAW264.7 macrophages and that the mechanisms involved downregulation of the FoxO3a/autophagy signaling pathway.
Project description:Alkaloids 1?10 were isolated from the aerial parts of Tetrastigma hemsleyanum (APTH) and obtained from species of the genus Tetrastigma for the first time. The chemical structures of the isolated compounds were identified by NMR, UV, and MS analyses. Their anti-inflammatory activities were investigated by measuring nitric oxide (NO) production in lipopolysaccharide (LPS)-induced RAW264.7 macrophages. Among all the isolates, compounds 6, 7 and 10 showed potent inhibitory activity against LPS-stimulated NO production in RAW264.7 cells (IC50: 31.9, 25.2 and 6.3 ?M, respectively). Furthermore, APTH and S-(?)-trolline (10) inhibited induction of inflammatory cytokines or mediators such as interleukin-1? (IL-1?) and inducible nitric oxide synthase (iNOS) via suppression of nuclear factor ?B (NF-?B) translocation into the nucleus. In addition, 10 suppressed extracellular signal-regulated protein kinase 1/2 (ERK1/2) mitogen-activated protein kinase (MAPK) phosphorylation in a dose-dependent manner. These results conclusively demonstrated that compound 10 displays anti-inflammatory activity via suppression of NF-?B activation and the ERK-MAPK signaling pathway in LPS-stimulated RAW264.7 cells.
Project description:Apios americana, a leguminous plant, is used as food in some countries. Although the biological activities of Apios extract have been reported, there have been no reports about the anti-inflammatory mechanism of lupinalbin A on the RAW264.7 cells. In this study, we investigated the anti-inflammatory effect of A. americana lupinalbin A on lipopolysaccharide (LPS)-treated RAW264.7 cells. Lupinalbin A significantly inhibited nitric oxide production and inducible nitric oxide synthase expression in LPS-treated RAW264.7 cells. The expression of cytokines, including interleukin-6, tumor necrosis factor-?, and chemokine of monocyte chemoattractant protein, was reduced under lupinalbin A exposure in LPS-treated RAW264.7 cells. In addition, lupinalbin A significantly decreased LPS-induced interferon (IFN)-? production and STAT1 protein levels in RAW264.7 cells. Taken together, these results suggest that A. americana lupinalbin A exerts anti-inflammatory effects via the inhibition of pro-inflammatory cytokines and blocking of IFN-?/STAT1 pathway activation.
Project description:Recent studies confirm that chronic low-grade inflammation is closely associated with metabolic syndromes, and anti-inflammatory therapy is a potential approach for treating cardiovascular diseases and type 2 diabetes. Accumulating evidence suggests that GPR120 activation is a feasible solution to ameliorating chronic inflammation and improving glucose metabolism. In this study we investigated whether ginsenoside Rb2 (Rb2), which exhibited regulatory activities in glucose and lipid metabolism, affected GPR120 expression in lipopolysaccharide (LPS)-activated mouse macrophage RAW264.7 cells, and examined the contribution of GPR120 activation to reducing the LPS-induced inflammatory response. LPS (100 ng/mL) activated the macrophages, resulting in dramatic increases in TNF-?, IL-6, IL-1? and NO production. Treatment with a ?-3 fatty acid ?-linolenic acid (ALA, 50 ?mol/L) produced moderate reduction in LPS-stimulated inflammatory cytokines and NO production (TNF-? and IL-6 were decreased by 46% and 42%, respectively). Pre-incubation with Rb2 (1 or 10 ?mol/L) for 12 h before ALA treatment dramatically amplified the inhibitory effects of ALA (TNF-? and IL-6 were decreased by 74% and 86%, respectively). Compared to the treatment with ALA alone, pre-incubation with Rb2 resulted in a more prominent reduction in LPS-stimulated expression of iNOS and COX-2 and LPS-stimulated IKK/NF-?B phosphorylation and MAPK pathway activation. Rb2 (0.1-100 ?mol/L) dose- and time-dependently increased both mRNA and protein expression of GPR120 in RAW264.7 cells, but treatment with Rb2 alone did not exert anti-inflammatory effect in LPS-activated RAW264.7 cells. In RAW264.7 cells transfected with GPR120 shRNA, the ameliorating effects of Rb2 on LPS-induced inflammation were abolished. In conclusion, Rb2 exerts anti-inflammatory effect in LPS-stimulated mouse macrophage RAW264.7 cells in vitro by increasing GPR120 expression and subsequently enhancing ?-3 fatty acid-induced GPR120 activation.
Project description:BACKGROUND:The uteruses of most dairy cattle are easily infected by bacteria, especially gram-negative bacteria, following parturition. Macrophages are important cells of the immune system and play a critical?role?in?the?inflammatory?response. In addition, cortisol levels become significantly increased due to the stress of parturition in dairy cattle, and cortisol is among the most widely used and effective therapies for many inflammatory diseases. In this study, we assessed the anti-inflammatory effects and potential molecular mechanisms of cortisol using a Lipopolysaccharide (LPS)-induced RAW264.7 macrophage cell line. RESULTS:Cortisol significantly suppressed the production of prostaglandin E2 (PGE2) and decreased the gene and protein expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) in a dose-dependent manner. Moreover, cortisol inhibited the mRNA expression of pro-inflammatory cytokines including tumor necrosis factor alpha (TNF?), interleukin-1? (IL-1?), and interleukin-6 (IL-6) and decreased IL-1? secretion in an LPS-treated RAW264.7 macrophage cell line. Moreover, we found that cortisol suppressed nuclear factor-kappa B (NF-?B) signaling in RAW264.7 macrophages stimulated with LPS. This suppression was mediated by the inhibition of I?B? degradation and NF-?B p65 phosphorylation. In addition, cortisol also suppressed the phosphorylation of mitogen-activated protein kinases (MAPK) such as extracellular signal-regulated kinase (ERK1/2), p38 MAPK, and c-Jun N-terminal kinase/stress-activated protein kinase (JNK). CONCLUSIONS:These results suggest that high cortisol levels can attenuate LPS-induced inflammatory responses in the RAW264.7 macrophage cell line by regulating the NF-?B and MAPK signaling pathways.
Project description:Inflammation is a complex physiological process that poses a serious threat to people's health. However, the potential molecular mechanisms of inflammation are still not clear. Moreover, there is lack of effective anti-inflammatory drugs that meet the clinical requirement. Procyanidin A1 (PCA1) is a monomer component isolated from Procyanidin and shows various pharmacological activities. This study further demonstrated the regulatory role of PCA1 on lipopolysaccharide (LPS)-stimulated inflammatory response and oxidative stress in RAW264.7 cells. Our data showed that PCA1 dramatically attenuated the production of pro-inflammatory cytokines such as NO, iNOS, IL-6, and TNF-α in RAW264.7 cells administrated with LPS. PCA1 blocked IκB-α degradation, inhibited IKKα/β and IκBα phosphorylation, and suppressed nuclear translocation of p65 in RAW264.7 cells induced by LPS. PCA1 also suppressed the phosphorylation of JNK1/2, p38, and ERK1/2 in LPS-stimulated RAW264.7 cells. In addition, PCA1 increased the expression of HO-1, reduced the expression of Keap1, and promoted Nrf2 into the nuclear in LPS-stimulated RAW264.7 cells. Cellular thermal shift assay indicated that PCA1 bond to TLR4. Meanwhile, PCA1 inhibited the production of intracellular ROS and alleviated the depletion of mitochondrial membrane potential in vitro. Collectively, our data indicated that PCA1 exhibited a significant anti-inflammatory effect, suggesting that it is a potential agent for the treatment of inflammatory diseases.
Project description:Mulberry fruit (Morus alba L.) contains abundant bioactive compounds, including anthocyanins and flavonols, and has been reported to possess potent beneficial properties including anticancer, antidiabetic, and anti-oxidant effects. High hydrostatic pressure (HHP) processing, a nonthermal food processing technology, is suitable for the extraction of bioactive compounds from plants. Nevertheless, the anti-inflammatory effects of HHP extract of mulberry fruit (HM) in RAW264.7 cells remain unclear. The present study aimed to investigate the anti-inflammatory effects of HM on lipopolysaccharide (LPS)-induced inflammation in vitro. RAW264.7 cells were treated with various concentrations (0.1-1 ?g/mL) of HM in the presence or absence of LPS. HM inhibited the inflammatory mediator, nitric oxide (NO) release, and mRNA expression of nitric oxide synthase 2 (NOS2) in LPS-induced RAW264.7 cells. In addition, HM suppressed both mRNA and protein expressions of prostaglandin-endoperoxide synthase 2 (PTGS2). Moreover, it reduced the LPS-induced secretion of proinflammatory cytokines such as interleukin (IL)-6 and tumor necrosis factor (TNF)-?. These results revealed that HM exerts anti-inflammatory effects by inhibiting several mediators and cytokines involved in the inflammatory process.
Project description:Inflammation is a common pathogenesis in many diseases. Salvia miltiorrhiza Bunge (Danshen), a traditional Chinese medicine, has been considered to have good anti-inflammatory effects. In the present study, we investigated the anti-inflammatory effect of diethyl blechnic (DB), a novel compound isolated from Danshen, and its possible mechanisms in lipopolysaccharide (LPS)-induced RAW264.7 macrophages. The results showed that DB can inhibit the LPS-induced pro-inflammatory cytokines release of prostaglandin E2 (PGE2) and mRNA expression of TNF-?, IL-6, and IL-1?. In addition, the results of the flow cytometry assay and the fluorometric intracellular ROS kit assay indicated that DB reduced the generation of ROS in LPS-stimualted RAW264.7 cells. DB reversed the LPS-induced loss of the mitochondrial membrane potential (MMP). Furthermore, DB suppressed the LPS-stimulated increased expression of Toll-like receptor 4 (TLR4), myeloid differential protein-88 (MyD88) and phosphorylation of TAK1, PI3K, and AKT. DB promoted NF-E2-related factor 2 (Nrf2) into the nucleus, increased the expression of heme oxygenase-1 (HO-1) and NAD(P)H dehydrogenase [quinone] 1 (NQO1) and reduced the expression of Keap1. In summary, DB may inhibit LPS-induced inflammation, which mainly occurs through TLR4/MyD88 and oxidative stress signaling pathways in RAW264.7 cells.