Ethanol extract of Chondracanthus tenellus(Harvey)Hommersand attenuates lipopolysaccharide-induced inflammatory and oxidative response by blocking the NF-κB,MAPKs,and PI3K/Akt signaling pathways

Objective:To investigate whether the ethanol extract of Chondracanthus tenellus(Harvey)Hommersand,a type of red algae,could exhibit anti-inflammatory potential in lipopolysaccharide(LPS)-stimulated macrophages.Methods:The ethanol extract of Chondracanthus tenellus was applied to 100 ng/mL LPS-stimulated RAW 264.7 cells,and cell viability,phagocytic ability,levels of pro-inflammatory factors,and the production of reactive oxygen species were measured.To identify the underlying mechanism of the ethanol extract of Chondracanthus tenellus,the expression of inflammation-regulated genes was estimated.Results:The ethanol extract of Chondracanthus tenellus had no cytotoxic effect at concentrations below 300μg/mL,and reduced the LPS-induced production of inflammatory mediators including nitric oxide(NO)and prostaglandin E2.Furthermore,the extract markedly suppressed the expression of inducible NO synthase and cyclooxygenase-2,as well as the production of reactive oxygen species.The LPS-induced up-regulation of pro-inflammatory cytokines was attenuated by treatment with the ethanol extract of Chondracanthus tenellus,reducing their extracellular secretion.The Chondracanthus tenellus extract also inhibited LPS-mediated activation of nuclear factor-kappa B(NF-κB).In addition,the phosphorylation of mitogen activated protein kinases(MAPKs)and phosphatidylinositol 3 kinase(PI3K)/Akt was markedly increased by LPS,which was significantly abolished by the Chondracanthus tenellus extract.Conclusions:Our findings indicate that the ethanol extract of Chondracanthus tenellus exhibited potential anti-inflammatory and antioxidant effects through downregulating the NF-κB,MAPKs,and PI3K/Akt signaling pathways in LPS stimulated RAW 264.7 macrophages.tenellus extract.Conclusions:Our findings indicate that the ethanol extract of Chondracanthus tenellus exhibited potential anti-inflammatory and antioxidant effects through downregulating the NF-κB,MAPKs,and PI3K/Akt signaling pathways in LPS stimulated RAW 264.7 macrophages.

Ethanol extract of Kalopanax septemlobus leaf inhibits HepG2 human hepatocellular carcinoma cell proliferation via inducing cell cycle arrest at G_1 phase

Objective:To investigate the effects of an ethanol extract of Kalopanax septemlobus(Thunb.)Koidz.leaf(EEKS) on cell proliferation in human hepatocellular carcinoma cells and its mechanisms of action.Methods:Cells were treated with EEKS and subsequently analyzed for cell proliferation and flow cytometry analysis.Expressions of cell cycle regulators were determined by reverse transcriptase polymerase chain reaction analysis and Western blotting,and activation of eyclin-associaled kinases studied using kinase assays.Results:The EEKS suppressed cell proliferation in both HepG2 and Hep3 B cells,but showed a more sensitive anli-proliferative activity in HepG2 cells.Flow cytometry analysis revealed an association between the growth inhibitory effect of EEKS and with G_1 phase cell cycle arrest in HepG2 cells,along with the dephosphorylation of retinoblastoma protein(pRB) and enhanced binding of pRB with the E2 F transcription factor family proteins.Treatment with EEKS also increased the expression of cyclin-dependent kinase(CDK) inhibitors,such as p21WAF1/CIP1 and p27KIP1.without any noticeable changes in G_1 cyclins and CDKs(except for a slight decrease in CDK4).Treatment of HepG2 cells with EEKS also increased the binding of p21 and p27 with CDK4 and CDK6.which was paralleled by a marked decrease in the cyclin D- and cyclin E-associated kinase activities.Conclusions:Overall,our findings suggest that EEKS may be an effective treatment for liver cancer through suppression of cancer cell proliferation via G_1,cell cycle arrest Further studies arc required to identify the active compounds in EEKS.

Ethanol extracts of Hizikia fusiforme induce apoptosis in human prostate cancer PC3 cells via modulating a ROS-dependent pathway

Objective:To investigate whether ethanol extracts of Hizikia fusiforme could induce apoptosis in human prostate cancer PC3 cells.Methods:Cell viability was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide.Apoptosis and mitochondrial membrane potential(MMP)were measured using flow cytometry in PC3 cells.DNA damage was assessed by nuclear staining and DNA fragmentation assay.Expressions of apoptosis-associated proteins were determined by Western blotting assays.Activities of caspase-3,-8,and-9 were determined by colorimetric assay.Moreover,intracellular reactive oxygen species(ROS)generation was detected using a flow cytometer and fluorescence microscope.Results:Treatment of PC3 cells with ethanol extracts of Hizikia fusiforme inhibited proliferation,which was associated with induction of apoptosis,and accompanied by increased expression of Fas,Fas-ligand(Fas L),Bax and t Bid,and decreased expression of Bcl-2.In addition,ethanol extracts of Hizikia fusiforme reduced c-Flip expression and activated caspase-8,-9 and-3,resulting in an increase in poly(ADP-ribose)polymerase(PARP)cleavage.However,in the presence of a pan-caspase inhibitor,ethanol extracts of Hizikia fusiforme-mediated growth inhibition and apoptosis were significantly attenuated.Ethanol extracts of Hizikia fusiforme also destroyed the integrity of mitochondria due to the loss of MMP,leading to cytosolic release of cytochrome c.Moreover,the levels of ROS were markedly increased by treatment with ethanol extracts of Hizikia fusiforme,which was significantly suppressed by the ROS scavenger N-acetyl-L-cysteine.Further investigation of whether ethanol extracts of Hizikia fusiforme-induced apoptosis was related to the generation of ROS was conducted and the results showed that N-acetyl-L-cysteine fully blocked ethanol extracts of Hizikia fusiforme-induced apoptotic events including loss of MMP,activation of caspase-3,the cytosolic release of cytochrome c and cytotoxicity.Conclusions:Ethanol extracts of Hizikia fusiforme have chemopreventive potential via induction of ROS-dependent apoptosis.Therefore,ethanol extracts of Hizikia fusiforme may be useful for developing effective and selective natural sources to inhibit cancer cell proliferation.

Immunostimulatory effect of ethanol extract of Chondracanthus tenellus in RAW 264.7 macrophages in vitro

Objective:To investigate whether ethanol extracts of Chondracanthus tenellus(EECT)could improve immunomodulatory property of murine monocyte/macrophage RAW 264.7 cells.Methods:Cell viability,phagocytic ability,and nitric oxide were measured.The levels of prostaglandin E2 and cytokines were determined using enzyme-linked immunosorbent assay kits.Expression of immunoregulatory response protein was detected by Western blotting assay.Results:As the concentration of EECT increased,the morphology of the cells changed to a typical active macrophage shape,and the phagocytic activity increased significantly.EECT also effectively enhanced the production and secretion of immunomodulatory mediators,such as nitric oxide and prostaglandin E2,and cytokines.In addition,compared with the control group,EECT markedly stimulated the expression of Toll-like receptor 4(TLR4)and myeloid differentiation factor 88,one of the TLR4 adapter molecules.Furthermore,EECT promoted the nucleus translocation of nuclear factor-kappa B(NF-κB)by increasing the phosphorylation and degradation of the inhibitor of NF-κB-α,indicating activation of the NF-κB signaling pathway.Meanwhile,similar trends were found in cells treated with lipopolysaccharide as a positive control.Conclusions:Taken together,the results indicate that EECT has an immunomodulatory effect by increasing the production of immunomodulatory mediators and cytokines through activation of the TLR4/NF-κB signaling pathway.EECT could be used as a potential candidate for medication or dietary supplements to increase immune activity.

Effect of Autologus Platelet-Rich Plasma on IL-6, MMP-3 and MCP-1 Expression in Synoviocytes from Osteoarthritic Patients Knees

Nowadays, some studies reported promising results of platelet-rich plasma (PRP) for the treatment of osteoarthritis (OA). However, the effects of PRP on prevention of osteoarthritis in knee joints have been debated. The present study investigated the effects of PRP on osteoarthritis related inflammatory cytokines expressed in fibroblast-like synoviocytes (FLS) from osteoarthritic knees. The synovial tissues were harvested from eight osteoarthritic patients who had undertaken total knee arthroplasties (TKAs) and cultured in DMEM containing 10% FBS. Platelet-rich plasma releasate (PRPr) was made by clotting or activation of PRP by citrate. The levels of PDGF-AA and VEGF in PRPr and whole blood were measured with ELISA method. The FLS were isolated and cultured from osteoarthritic knees. The IL-1β stimulated FLS were cultivated with three different conditions (none, 1% and 10% of PRP). To determine the expression of IL-6, MMP-3, and MCP-1, we used reverse transcriptase polymerase chain reaction (RT-PCR). The concentrations of platelet count in PRP were about 7 to 9 times higher than those of whole blood. The levels of PDGF-AA in PRPr were approximately 3 to 4 times higher than those of whole blood. The levels of VEGF in PRPr were also significantly 7 to 18 times higher than those of whole blood. Without induction of the FLS with IL-1β, 1% or 10% PRPr did not reduce expressions of inflammatory proteins (MMP-3, MCP-1), except for IL-6. However, with induction of the FLS with IL-1β, both concentrations (1% and 10%) of PRPr reduced significantly all inflammatory protein expressions (IL-6, MMP-3, MCP-1). PRPr diminished inflammatory IL-1β-mediated effects on human osteoarthritic fibroblast-like synoviocytes. These results suggest that platelet-rich plasma can be a good therapeutic option for the treatment of osteoarthritis.