Ferulic acid inhibits H2O2-induced oxidative stress and inflammation in rat vascular smooth muscle cells via inhibition of the NADPH oxidase and NF-κB pathway

Ferulic acid (FA) is a dietary phenolic acid and has a wide range of therapeutic effects, including anti-aging, antitumor activity and antihypertensive effects. The aim of present study was to evaluate the inhibitory effects of FA on cell inflammation and oxidative stress against hydrogen peroxide (H₂O₂)-induced injury in rat vascular smooth muscle cells (VSMCs) in vitro. VSMCs were pretreated with FA 2 h before H₂O₂ incubation. The results suggested that FA inhibited H₂O₂-induced cell injury by reducing the MDA and increasing the SOD activity and GSH content. In rat VSMCs exposed to H₂O₂, FA increased the cell viability and restored the mitochondrial membrane depolarization. The level of ROS generation was reduced by pretreatment with FA through inhibiting the expression of NADPH oxidase and down-regulating MAPK and AKT pathways. We found that H₂O₂ stimulated the production of IL-6, IL-1β, TNF-α and NO, which could be reduced by pretreatment with FA through inhibiting the p-NF-κB as well as the iNOS expression. In conclusion, our results show that FA may serve as a novel drug in the treatment of these pathologies by inhibiting NADPH oxidase and NF-κB and subsequently decreasing VSMC oxidative stress and inflammation. These suggest that the inhibitory effect of FA on VSMC inflammation and oxidative stress is partially attributed to depressing NADPH and NF-κB expressions in VSMCs, decreasing the ROS production and reducing apoptosis of VSMCs.     

2-Hydroxycircumdatin C inhibits LPS-induced BV2 cells inflammatory response via down-regulation of TLR4-mediated NF-κB/MAPK and JAK2/STAT3 pathways

2-Hydroxycircumdatin C (2-HCC) is a circumdatin-type alkaloid isolated from a coral-associated fungus Aspergillus ochraceus LZDX-32-15. In the present study, we aimed to assess the neuroprotective effects of 2-HCC on the microglia-mediated inflammatory response as well as underlining molecular mechanisms. 2-HCC could significantly down-regulate the overproduction of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) induced by lipopolysaccharide (LPS) both in BV2 cells and primary microglial cells without affecting cell viability. In addition, 2-HCC exerted obvious neuroprotective effects against inflammatory injury in neurons when cocultured with LPS-induced microglia. Mechanism investigation indicated that the anti-inflammatory effect of 2-HCC involved the inhibition of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2) and alleviation of the LPS-induced TLR4-NF-κB/MAPK pathway. Furthermore, 2-HCC treatment attenuated LPS-induced activation of the JAK2/STAT3 pathway. In conclusion, these results indicated that the anti-inflammatory and neuroprotective properties of 2-HCC, at least partially, depended upon TLR4-NF-κB/MAPK and JAK2/STAT3 signaling pathways.     

Isotrifoliol inhibits pro-inflammatory mediators by suppression of TLR/NF-κB and TLR/MAPK signaling in LPS-induced RAW264.7 cells

Soybeans, produced by Glycine max (L.) Merr., contain high levels of isoflavones, such as genistein and daidzein. However, soy leaves contain more diverse and abundant flavonol glycosides and coumestans, as compared to the soybean. This study investigated the anti-inflammatory effects of the major coumestans present in soy leaf (coumestrol, isotrifoliol, and phaseol) in lipopolysaccharide (LPS)-induced RAW264.7 cells. Coumestans significantly reduced LPS-induced nitric oxide (NO), prostaglandin E2 (PGE₂), and reactive oxygen species (ROS) production; isotrifoliol had the most potent anti-inflammatory activity. Isotrifoliol reduced LPS-mediated induction of mRNA expression of inducible nitric-oxide synthase (iNOS), cyclooxygenase-2 (COX-2), interleukin (IL)-1β, IL-6, tumor necrosis factor alpha (TNFα), and chemokines, such as chemokine (C-C motif) ligand (CCL) 2, CCL3, and CCL4. Isotrifoliol prevented NF-κB p65 subunit activation by reducing the phosphorylation and degradation of the inhibitor of NF-κB. And isotrifoliol significantly suppressed phosphorylation of the extracellular signal-regulated protein kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK). Furthermore, isotrifoliol suppressed LPS-induced Toll-like Receptor (TLR) signaling pathway, including mRNA expression of TNF receptor associated factor 6, transforming growth factor beta-activated kinase 1 (TAK1), TAK1 binding protein 2 (TAB2), and TAB3. These results demonstrate that isotrifoliol exerts an anti-inflammatory effect by suppressing the expression of inflammatory mediators via inhibition of TLR/NF-κB and TLR/MAPK signaling in LPS-induced RAW264.7 macrophages. Therefore, isotrifoliol can be used as an anti-inflammatory agent, and coumestan-rich soy leaf extracts may provide a useful dietary supplement.     

Anti-inflammatory effects of sophocarpine in LPS-induced RAW 264.7 cells via NF-κB and MAPKs signaling pathways

Sophocarpine, a tetracyclic quinolizidine alkaloid, is one of the most abundant active ingredients in Sophora alopecuroides L. Our previous studies have showed that sophocarpine exerts anti-inflammatory activity in animal models. In the present study, anti-inflammatory mechanisms of sophocarpine were investigated in lipopolysaccharide (LPS)-induced responses in RAW 264.7 cells. Furthermore, the cytotoxicity of sophocarpine was tested. The results indicated that sophocarpine could increase the LDH level and inhibit cell viability up to 800μg/ml, and which was far higher than that of the plasma concentration of sophocarpine in clinical effective dosage. The results also demonstrated that sophocarpine (50 and 100μg/ml) suppressed LPS-stimulated NO production and pro-inflammatory cytokines secretion, including tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6). These were associated with the decrease of the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Furthermore, sophocarpine inhibited LPS-mediated nuclear factor-κB (NF-κB) activation via the prevention of inhibitor κB (IκB) phosphorylation. Sophocarpine had no effect on the LPS-induced phosphorylation of extracellular signal-regulated kinase 1/2 (Erk1/2), whereas it attenuated the phosphorylation of p38 mitogen-activated protein (MAP) kinase and c-Jun NH(2)-terminal kinase (JNK). Our data suggested that sophocarpine exerted anti-inflammatory activity in vitro, and it might attribute to the inhibition of iNOS and COX-2 expressions via down-regulation of the JNK and p38 MAP kinase signal pathways and inhibition of NF-κB activation.     

Neurotropin inhibits neuroinflammation via suppressing NF-κB and MAPKs signaling pathways in lipopolysaccharide-stimulated BV2 cells

Neurotropin (NTP) is a widely used drug in China and Japan mainly for the treatment of chronic pain and peripheral inflammation. Nevertheless, the effects of NTP on neuroinflammation have not been explored. In this study, we investigated the anti-inflammatory effects of NTP in lipopolysaccharide (LPS)-stimulated BV-2 microglial cells and its underlying mechanisms. BV-2 cells were pretreated with NTP for 12 h before exposure to LPS. The expression of pro-inflammatory cytokines (TNF-α and IL-6) were detected by RT-PCR and EILSA at mRNA and protein levels, respectively. Western blotting was conducted to measure the protein levels of major genes in MAPKs and NF-κB signaling pathways. Results demonstrated that NTP could attenuate the production of pro-inflammatory cytokines. Furthermore, NTP inhibited the activation of NF-κB signaling by decreasing the translocation of NF-κB p65 to the nucleus and suppressed the MAPKs signaling pathway via inhibition of the phosphorylation of p38, ERK and JNK. Taken together, these findings suggest that neurotropin exerts anti-inflammatory effects by suppressing the production of pro-inflammatory mediators via inhibition of NF-κB and MAPKs signaling pathways in LPS-stimulated BV-2 cells.     

Desoxyrhapontigenin,a potent anti-inflammatory phytochemical,inhibits LPS-induced inflammatory responses via suppressing NF-κB and MAPK pathways in RAW 264.7 cells

This study investigates the anti-inflammatory effects of a stilbene compound, desoxyrhapontigenin, which was isolated from Rheum undulatum. To determine the anti-inflammatory effects of this compound, lipopolysaccharide (LPS)-induced RAW 264.7 macrophages were treated with different concentrations of six stilbene derivatives. The results indicated that compared with other stilbene compounds, desoxyrhapontigenin (at 10, 30 and 50 μM concentrations) significantly inhibited nitric oxide (NO) production, nuclear factor kappa B (NF-κB) activation, the protein expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression. Therefore, the anti-inflammatory mechanism of desoxyrhapontigenin was investigated in detail. The results of this investigation demonstrated that desoxyrhapontigenin suppressed not only LPS-stimulated pro-inflammatory cytokine secretions, including the secretions of tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6), but also PGE₂ release. As assayed by electrophoretic mobility shift assays (EMSAs), desoxyrhapontigenin also produced the dose-dependent inhibition of the LPS-induced activation of NF-κB and AP-1. Moreover, desoxyrhapontigenin inhibited the protein expression of myeloid differentiation primary response gene 88 (MyD88), IκB kinase (IKK) phosphorylation and the degradation of IκBα. Activations of p-JNK1 and p-Akt were also significantly inhibited, and phosphorylation of p38 and ERK was down-regulated. A further study revealed that desoxyrhapontigenin (5 and 25 mg/kg, i.p.) reduced paw swelling in carrageenan-induced acute inflammation model in vivo. On the whole, these results indicate that desoxyrhapontigenin showed anti-inflammatory properties by the inhibition of iNOS and COX-2 expression via the down-regulation of the MAPK signaling pathways and the inhibition of NF-κB and Akt activation.     

The mineralocorticoid receptor-p38MAPK-NFκB or ERK-Sp1 signal pathways mediate aldosterone-stimulated inflammatory and profibrotic responses in rat vascular smooth muscle cells

Aim:

To explore the signalling pathways involved in aldosterone-induced inflammation and fibrosis in rat vascular smooth muscle cells (VSMCs).

Methods:

Using Western blotting and real-time RT-PCR, we investigated the effects of aldosterone on the expression of cyclooxygenase-2 (Cox-2) and IL-6, two important proinflammatory factors, and TGFβ1, a critical profibrotic factor, in VSMCs.

Results:

Aldosterone treatment significantly increased the expression of Cox-2 and IL-6 and activation of p38MAPK and NF-κB. The expression of both Cox-2 and IL-6 could be blocked by the mineralocorticoid receptor (MR) antagonist spironolactone and the p38MAPK inhibitor SB203580. Also, the rapid phosphorylation of p38MAPK could be suppressed by SB203580 but not by spironolactone, implicating in nongenomic effects of aldosterone. Similar to SB203580 and spironolactone, the NF-κB inhibitor α-p-tosyl-L-lysine chloromethyl ketone (TLCK) markedly attenuated expression of Cox-2, indicating that MR, p38MAPK and NF-κB are associated with aldosterone-induced inflammatory responses. Furthermore, aldosterone enhanced expression of TGFβ1 in rat VSMCs. This result may be related to activation of the MR/ERK-Sp1 signalling pathway because PD98059, an ERK1/2 inhibitor, significantly blocked the rapid phosphorylation of ERK1/2 and function of Sp1 and led to reduced expression of TGFβ1. Spironolactone was also shown to significantly inhibit TGFβ1 and Sp1 expression but not ERK1/2 phosphorylation.

Conclusion:

These results suggest that aldosterone-induced inflammatory responses and fibrotic responses may be mediated by the MR/p38MAPK-NF-κB pathways and the MR/ERK-Sp1 pathways in VSMCs, respectively.     

The procyanidin trimer C1 inhibits LPS-induced MAPK and NF-κB signaling through TLR4 in macrophages

Natural products and dietary components rich in polyphenols have been shown to reduce inflammation; however, the molecular mechanisms underlying this anti-inflammatory activity are not completely characterized, and many features remain to be elucidated. This research was carried out to clarify the potential role of procyanidin trimer C1 in the anti-inflammatory effect of polyphenols. Procyanidin C1 inhibited inducible nitric oxide synthase-mediated nitric oxide production and the release of pro-inflammatory cytokines (interleukin-6 and tumor necrosis factor-α) in lipopolysaccharide (LPS)-induced macrophages. Treatment with procyanidin C1 resulted in a significant decrease in prostaglandin E₂ and cyclooxygenase-2 levels, as well as the expression of cell surface molecules (CD80, CD86, and MHC class II), which was induced by LPS. Furthermore, our data demonstrated that the anti-inflammatory effect of procyanidin C1 occurs through inhibition of mitogen-activated protein kinase (p38 and c-Jun N-terminal kinase) and nuclear factor-κB signaling pathways. These 2 factors play a major role in controlling inflammation, through toll-like receptor 4, suggesting that procyanidin C1 plays a potent role in promoting anti-inflammatory activity in macrophages. These results represent a novel and effective therapeutic intervention for the treatment of inflammatory disease.     

Vaspin prevents TNF-α-induced intracellular adhesion molecule-1 via inhibiting reactive oxygen species-dependent NF-κB and PKCθ activation in cultured rat vascular smooth muscle cells

Vaspin (visceral adipose tissue-derived serine protease inhibitor) was recently identified as a novel adipocytokine with insulin-sensitizing effects. We hypothesized that vaspin could play a role in vascular inflammation. To test the hypothesis, we investigated the effects of vaspin on TNF-α-stimulated vascular smooth muscle cells (SMCs) focusing on inflammatory signal transduction. Vascular SMCs from mesenteric artery of male Wistar rats were treated with TNF-α (5–10 ng/ml, 20 min–6 h) in the absence or presence of vaspin (1–300 ng/ml, pretreatment for 24 h). Western blotting was performed to analyze the cellular signal. Reactive oxygen species (ROS) generation was fluorometrically measured using 2′,7′-diclorofluorescein diacetate. Vaspin alone treatment had no effect on vascular SMCs morphology and cellular signal. Vaspin significantly decreased the TNF-α-induced monocyte adhesion to SMCs. Vaspin significantly inhibited the protein expression of intracellular adhesion molecule (ICAM)-1 and the phosphorylation of NF-κB and protein kinase C (PKC)θ induced by TNF-α. Both of NF-κB and novel PKC inhibitors significantly attenuated the TNF-α-induced ICAM-1 expression. Moreover, vaspin inhibited TNF-α-induced ROS generation. An anti-oxidant, N-acetyl-l-cysteine blocked the TNF-α-induced activation of NF-κB, PKCθ and expression of ICAM-1. The present results demonstrated for the first time that vaspin inhibits TNF-α-induced expression of ICAM-1 via preventing the ROS generation and subsequent activation of NF-κB and PKCθ. Consequently, vaspin could play inhibitory roles on inflammatory states of vascular SMCs.     

Inhibition of TNF-α-induced adhesion molecule expression by diosgenin in mouse vascular smooth muscle cells via downregulation of the MAPK, Akt and NF-κB signaling pathways

Atherosclerosis is a chronic inflammatory disease and the expression of adhesion molecules on vascular smooth muscle cells (VSMCs) contributes to the progress of the disease. Diosgenin, a precursor of steroid hormones, has been shown to have a variety of biological activities including anti-inflammatory activity; however, its molecular mechanisms are poorly understood. This study examined the effect of diosgenin on the expression of adhesion molecules induced by TNF-α in cultured mouse VSMC cell line, MOVAS-1. Preincubation of VSMCs for 2h with diosgenin (0.1-10 μM) dose-dependently inhibited TNF-α-induced adhesion of THP-1 monocytic cells and mRNA and protein expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1). Diosgenin abrogated TNF-α induced production of intracellular reactive oxygen species (ROS) and phosphorylation of p38, ERK, JNK and Akt. Diosgenin was also shown to inhibit NK-κB activation induced by TNF-α. Furthermore, diosgenin inhibited TNF-α-induced IκB kinase activation, subsequent degradation of IκBα, and nuclear translocation of NF-κB. Our results indicate that diosgenin inhibits the adhesive capacity of VSMC and the TNF-α-mediated induction of ICAM-1 and VCAM-1 in VSMC by inhibiting the MAPK/Akt/NF-κB signaling pathway and ROS production, which may explain the ability of diosgenin to suppress inflammation within the atherosclerotic lesion and modulate immune response.     

Baicalin prevents LPS-induced activation of TLR4/NF-κB p65 pathway and inflammation in mice via inhibiting the expression of CD14

Previous studies have shown that baicalin,an active ingredient of the Chinese traditional medicine Huangqin,attenuates LPS-induced inflammation by inhibiting the activation of TLR4/NF-κBp65 pathway,but how it affects this pathway is unknown.It has been shown that CD14 binds directly to LPS and plays an important role in sensitizing the cells to minute quantities of LPS via chaperoning LPS molecules to the TLR4/MD-2 signaling complex.In the present study we investigated the role of CD14 in the anti-inflammatory effects of baicalin in vitro and in vivo.Exposure to LPS(1μg/mL)induced inflammatory responses in RAW264.7 cells,evidenced by marked increases in the expression of MHC II molecules and the secretion of NO and IL-6,and by activation of MyD88/NF-κB p65 signaling pathway,as well as the expression of CD14 and TLR4.These changes were dose-dependently attenuated by pretreatment baicalin(12.5–50μM),but not by baicalin post-treatment.In RAW264.7 cells without LPS stimulation,baicalin dose-dependently inhibit the protein and mRNA expression of CD14,but not TLR4.In RAW264.7 cells with CD14 knockdown,baicalin pretreatment did not prevent inflammatory responses and activation of MyD88/NF-κB p65 pathway induced by high concentrations(1000μg/mL)of LPS.Furthermore,baicalin pretreatment also inhibited the expression of CD14 and activation of MyD88/NF-κB p65 pathway in LPS-induced hepatocyte-derived HepG2 cells and intestinal epithelial-derived HT-29 cells.In mice with intraperitoneal injection of LPS and in DSS-induced UC mice,oral administration of baicalin exerted protective effects by inhibition of CD14 expression and inflammation.Taken together,we demonstrate that baicalin pretreatment prevents LPS-induced inflammation in RAW264.7 cells in CD14-dependent manner.This study supports the therapeutic use of baicalin in preventing the progression of LPS-induced inflammatory diseases.     

Inhibition of VCAM-1 expression on mouse vascular smooth muscle cells by lobastin via downregulation of p38, ERK 1/2 and NF-κB signaling pathways

Atherosclerosis is a chronic inflammatory disease, the progression of which is associated with the increased expression of cell adhesion molecules on vascular smooth muscle cells (VSMCs). Lobastin is a new pseudodepsidone isolated from Stereocaulon alpinum, Antarctic lichen, which is known to have antioxidant and antibacterial activities. However, the nature of the biological effects of lobastin still remains unclear. In the present study, we examine the effect of lobastin on the expression of vascular cell adhesion molecules (VCAM-1) induced by TNF-α in the cultured mouse VSMC cell line, MOVAS-1. Pretreatment of VSMCs for 2 h with lobastin (0.1–10 μg/ml) concentration-dependently inhibited TNF-α-induced protein expression of VCAM-1. Lobastin also inhibited TNF-α-induced production of intracellular reactive oxygen species (ROS). Lobastin abrogated TNF-α-induced phosphorylation of p38 and ERK 1/2, but not JNK, and also inhibited TNF-α-induced NK-κB activation. In addition, lobastin suppressed TNF-α-induced IκB kinase activation, subsequent degradation of IκBα and nuclear translocation of p65 NF-κB. Our results indicate that lobastin downregulates the TNF-α-mediated induction of VCAM-1 in VSMC by inhibiting the p38, ERK 1/2 and NF-κB signaling pathways and intracellular ROS generation. Thus, lobastin may be an important regulator of inflammation in the atherosclerotic lesion and a novel therapeutic drug for the treatment of atherosclerosis.     

Berberine suppresses LPS-induced inflammation through modulating Sirt1/NF-κB signaling pathway in RAW264.7 cells

Chronic inflammation is a major contributing factor in the pathogenesis of many diseases. Natural product berberine (BBR) exhibits potent anti-inflammatory effect in vitro and in vivo, while the underlying mechanisms remain elusive. Sirt1, a NAD⁺-dependent protein deacetylase, was recently found to play an important role in modulating the development and progression of inflammation. Thus, we speculate that Sirt1 might mediate the inhibitory effect of BBR on inflammation. In LPS-stimulated RAW264.7 macrophages, BBR treatment significantly downregulated the expression of proinflammatory cytokines such as MCP-1, IL-6 and TNF-α. Importantly, BBR potently reversed LPS-induced down-regulation of Sirt1. Consistently, the inhibitory effects of BBR on proinflammatory cytokines expression was largely abrogated by Sirt1 inhibition either by EX527, a Sirt1 inhibitor or Sirt1 siRNA. Further mechanistic studies revealed that BBR-induced inhibition of NF-κB is Sirt1-dependent, as either pharmacologically or genetically inactivating Sirt1 enhanced the IκΒα degradation, IKK phosphorylation, NF-κB p65 acetylation and DNA-binding activity. Taken together, our results provide the first evidence that BBR potently suppressed inflammatory responses in macrophages through inhibition of NF-κB signaling via Sirt1-dependent mechanisms.     

PPARα activator fenofibrate modulates angiotensin II-induced inflammatory responses in vascular smooth muscle cells via the TLR4-dependent signaling pathway

Angiotensin II (Ang II) is a crucial contributor to inflammatory processes involved in development and progression of atherosclerotic lesion. Toll-like receptor 4 (TLR4) signaling responsible for the initiation of inflammation also participates in pathogenesis of atherosclerosis. The protective effect of peroxisome proliferator-activated receptor α (PPARα) activators on atherosclerosis may be due to their impact on vascular inflammation, plaque instability and thrombosis. However, mechanisms underlying the inhibitory effects of PPARα activators on Ang II-induced vascular inflammation and the TLR4-dependent signaling pathway involved in vascular smooth muscle cells (VSMCs) remain unclear. The present study demonstrated that PPARα activator fenofibrate decreased Ang II-induced generation of pro-inflammatory mediators such as TLR4, MMP-9 and TNF-α, but enhanced production of anti-inflammatory molecules like PPARα and 6-keto-PGF_1α both in vivo and in vitro. Meanwhile, treatment of VSMCs with the TLR4 inhibitor or TLR4 siRNA showed that the inhibitory effects of fenofibrate on Ang II-induced inflammatory responses in VSMCs were dependent on TLR4. Furthermore, fenofibrate depressed Ang II-induced inflammatory responses in VSMCs by intervening the downstream effector molecules of the TLR4-dependent signaling pathway, including interferon-gamma inducible protein 10 (IP-10), protein kinases C (PKC) and nuclear factor κB (NF-κB). Thus, these findings provide the evidence for beneficial effects of PPARα activator fenofibrate to counter-regulate vascular inflammation induced by Ang II. More importantly, anti-inflammatory action of fenofibrate via interfering with the TLR4-dependent signaling pathway (TLR4/IP-10/PKC/NF-κB) works in concert to protect against atherosclerosis.     

4,7-Dimethoxy-5-methyl-1,3-benzodioxole from Antrodia camphorata inhibits LPS-induced inflammation via suppression of NF-κB and induction HO-1 in RAW264.7 cells

Several benzenoid compounds have been isolated from Antrodia camphorata are known to have excellent anti-inflammatory activity. In this study, we investigated the anti-inflammatory potential of 4,7-dimethoxy-5-methyl-1,3-benzodioxole (DMB), one of the major benzenoid compounds isolated from the mycelia of A. camphorata. DMB significantly decreased the LPS-induced production of pro-inflammatory molecules, such as nitric oxide (NO), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α) in RAW264.7 cells. In addition, DMB suppressed the protein levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in a dose dependent manner. Moreover, DMB significantly suppressed LPS-induced nuclear translocation of nuclear factor-κB (NF-κB), and this inhibition was found to be associated with decreases in the phosphorylation and degradation of its inhibitor, inhibitory κB-α (IκB-α). Moreover, we found that DMB markedly inhibited the protein expression level of Toll-like receptor 4 (TLR4). Furthermore, treatment with DMB significantly increased hemoxygenase-1 (HO-1) expression in RAW264.7 cells, which is further confirmed by hemin, a HO-1 enhancer, significantly attenuated the LPS-induced pro-inflammatory molecules and iNOS and TLR4 protein levels. Taken together, the present study suggests that DMB may have therapeutic potential for the treatment of inflammatory diseases.     

Renoprotective effects of Tilianin in diabetic rats through modulation of oxidative stress via Nrf2-Keap1 pathway and inflammation via TLR4/MAPK/NF-κB pathways

The present study was undertaken to assess the protective effects of Tilianin (TN) on type-2 diabetes-induced renal dysfunction in experimental rats. Diabetes was induced by injecting Nicotinamide (110 mg/kg) and streptozotocin (55 mg/kg) by i.p. and then the rats were treated with TN (10 and 20 mg/kg) daily by oral gavage for 28 days. TN treatment significantly decreases the BUN, creatinine, 24-hour urinary protein, urea, uric acid, and albumin protein levels. The protein of expression of Nrf2, NQO1, and HO-1 was augmented while the expression of Keap-1 decreased significantly. TN also reduces the oxidative/nitrosative status by lowering MDA content, NO, and MPO levels. TN exerted anti-inflammatory effects by suppressing TLR4/NF-κB/MAPK signaling cascades and inhibiting MyD88, TRAF6, IκBα, p38MAPK, JNK, and ERK2 in the diabetic rats. Histopathological findings supported the biochemical and molecular results. The results showed that TN modulated Nrf2-Keap1 and TLR4/MAPK/NF-κB signaling pathways and provided significant protection against diabetes-induced renal dysfunction.     

Neferine inhibits angiotensin Ⅱ-stimulated proliferation in vascular smooth muscle cells through heme oxygenase-1

Aim:

To explore the effect of neferine on angiotensin II (Ang II)-induced vascular smooth muscle cell (VSMC) proliferation.

Methods:

Human umbilical vein smooth muscle cells (HUVSMCs) were used. Cell proliferation was determined by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry analysis. Heme oxygenase (HO)-1 protein expression was tested by Western blot analysis. Extracellular signal-regulated protein kinase 1/2 (ERK1/2) activation was determined by using immunoblotting.

Results:

Pre-incubation of HUVSMCs with neferine (0.1, 0.5, 1.0, and 5.0 μmol/L) significantly inhibited Ang II-induced cell proliferation in a concentration-dependent manner and neferine 5.0 μmol/L increased HO-1 expression by 259% compared with control. The antiproliferative effect of neferine was significantly attenuated by coapplication of zinc protoporphyrin IX (ZnPP IX, an HO-1 inhibitor) with neferine. Ang II-enhanced ERK1/2 phosphorylation was markedly reversed by neferine. By inhibiting HO-1 activity with ZnPP IX, the inhibitive effect of neferine on ERK1/2 phosphorylation was significantly attenuated. Cobalt-protoporphyrin (CoPP), an HO-1 inducer, significantly decreased Ang II-induced ERK1/2 phosphorylation and inhibited Ang II-induced cell proliferation. The ERK1/2 pathway inhibitor PD98059 significantly blocked Ang II-enhanced ERK1/2 phosphorylation and inhibited cell proliferation.

Conclusion:

These findings suggest that neferine can inhibit Ang II-induced HUVSMC proliferation by upregulating HO-1, leading to the at least partial downregulation of ERK1/2 phosphorylation.     

Sulforaphane suppresses vascular adhesion molecule-1 expression in TNF-α-stimulated mouse vascular smooth muscle cells: involvement of the MAPK, NF-κB and AP-1 signaling pathways

Atherosclerosis is a long-term inflammatory disease of the arterial wall. Increased expression of the cell adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) is associated with increased proliferation of vascular smooth muscle cells (VSMCs), leading to increased neointima or atherosclerotic lesion formation. Therefore, the functional inhibition of adhesion molecules could be a critical therapeutic target of inflammatory disease. In the present study, we investigate the effect of sulforaphane on the expression of VCAM-1 induced by TNF-α in cultured mouse vascular smooth muscle cell lines. Pretreatment of VSMCs for 2h with sulforaphane (1-5μg/ml) dose-dependently inhibited TNF-α-induced adhesion of THP-1 monocytic cells and protein expression of VCAM-1. Sulforaphane also suppressed TNF-α-induced production of intracellular reactive oxygen species (ROS) and activation of p38, ERK and JNK. Furthermore, sulforaphane inhibited NK-κB and AP-1 activation induced by TNF-α. Sulforaphane inhibited TNF-α-induced ΙκΒ kinase activation, subsequent degradation of ΙκΒα and nuclear translocation of p65 NF-κB and decreased c-Jun and c-Fos protein level. This study suggests that sulforaphane inhibits the adhesive capacity of VSMC and downregulates the TNF-α-mediated induction of VCAM-1 in VSMC by inhibiting the MAPK, NF-κB and AP-1 signaling pathways and intracellular ROS production. Thus, sulforaphane may have beneficial effects to suppress inflammation within the atherosclerotic lesion.