A. Helen

Quercetin regulates oxidized LDL induced inflammatory changes in human PBMCs by modulating the TLR-NF-κB signaling pathway.

Toll-like receptors (TLRs) have been shown to play a pivotal role in both innate and adaptive immune responses. TLR family is the essential recognition and signaling component of mammalian host defense. Both genetic and biochemical data support a common signaling pathway that finally leads to the activation of NF-κB and induction of the cytokines and co-stimulatory molecules required for the activation of the adaptive immune response. The present study was designed to examine the involvement of TLR2 and TLR4 in the oxidized LDL induced inflammation in human PBMCs and the effect of flavonoid quercetin on TLR-NF-κB signaling mechanism. LDL was isolated from human plasma and oxidation of LDL was done by incubating with 10 μM CuSO₄ overnight at 37°C. The isolated human PBMCs in culture were used as the model system. 50 μg/ml ox-LDL treatment significantly up regulated TLR2 and TLR4 expression in isol human PBMCs after 24 h of culture and this was down regulated by quercetin at 25 μM concentration. ox-LDL caused a significant activation of NF-κB as evidenced by the detection of enhanced p65 subunit in nuclear extracts. Supplementation of quercetin significantly modulates the NF-κB p65 nuclear translocation. The cytokine IL-6 production was significantly increased in ox-LDL treated group and was decreased by quercetin treatment. Quercetin mediated reduction of TLR2 and TLR4 expression and the inhibition of nuclear translocation of NF-κB p65 in turn decreased the inflammatory enzymes like 5-LOX and COX and also decreased the mRNA expression of inducible enzymes like COX-2 and iNOS. Quercetin inhibited the ox-LDL induced TLR2 and TLR4 expression at mRNA level and modulated the TLR-NF-κB signaling pathway thereby inhibited the cytokine production and down regulated the activity of inflammatory enzymes thus have protective effect against the ox-LDL induced inflammation in PBMCs.

Anti-inflammatory and antioxidative effects of mucilage of Trigonella foenum graecum (Fenugreek) on adjuvant induced arthritic rats.

The aim of this study was to investigate anti-inflammatory and antioxidant effects of mucilage from fenugreek in adjuvant induced arthritis in rats. Arthritis was induced by intradermal injection of complete Freunds adjuvant into the right hind paw produce inflammation of the joint. The activities of inflammatory enzymes like cyclooxygenase, lipoxygenase and myeloperoxidase, and levels of nitrite and C-reactive protein were observed. Also oxidative stress was measured by analyzing the activity of catalase, superoxide dismutase, glutathione peroxidase and the levels of glutathione and vitamin C and lipid peroxidation. The blood parameters like ESR, total WBC, RBC and hemoglobin content was checked. Fenugreek mucilage exhibited maximum percentage of edema inhibition at a dose of 75 mg/kg on 21st day of adjuvant arthritis. The effect was higher than that of standard drug indomethacin. The activities of cyclooxygenase-2 and myeloperoxidase and concentration of thiobarbituric acid reactive substance (TBARS) were decreased and the activities of antioxidant enzymes, vitamins C and reduced glutathione level were increased on treatment with fenugreek mucilage. The increment in ESR and total WBC, reduction in RBC count and hemoglobin and aberrant changes to the C-reactive protein (CRP) levels observed in the arthritic animals were also found to be significantly restored in fenugreek mucilage treated rats. Histopathology of paw tissue showed decreased edema formation and cellular infiltration on supplementation with fenugreek mucilage. Thus the results demonstrated the potential beneficiary effect of fenugreek mucilage on adjuvant induced arthritis in rats.

Betulinic acid inhibits endotoxin‐stimulated phosphorylation cascade and pro‐inflammatory prostaglandin E2 production in human peripheral blood mononuclear cells

BACKGROUND AND PURPOSE Betulinic acid (BA) is a naturally occurring triterpenoid widely distributed throughout the plant kingdom. We previously reported that BA inhibits lipopolysaccharide (LPS)‐induced interleukin‐6 production through modulation of nuclear factor κB (NF‐κB) in human peripheral blood mononuclear cells (hPBMCs). This study attempted to identify other mechanisms through which BA modulates LPS signalling in mononuclear cells. The effects of BA on signalling pathways downstream were focused on in this study.

Betulinic acid isolated from Bacopa monniera (L.) Wettst suppresses lipopolysaccharide stimulated interleukin-6 production through modulation of nuclear factor-κB in peripheral blood mononuclear cells.

The purpose of this study was to investigate the anti-inflammatory function and mechanism(s) of action of an active component-betulinic acid isolated from Bacopa monniera. Betulinic acid, a pentacyclic triterpenoid markedly suppressed lipopolysaccharide (LPS) induced IL-6 production in blood mononuclear cells both in vivo and in vitro. Betulinic acid also prevented LPS induced nuclear translocation of p65 NF-kappaB in hPBMCs. LPS induced nuclear translocation of NF-kappaB and IL-6 production was also abolished by p38 and ERK MAPK inhibitors PD98059 and SB203580. Addition of each of these inhibitors to cell cultures along with betulinic acid caused significant downregulation of IL-6 production and inhibition of p65 NF-kappaB nuclear translocation. The inhibitory effect of both betulinic acid and the inhibitors was higher than that of cells treated with inhibitors alone. These results suggest that betulinic acid inhibited IL-6 production by preventing p65 NF-kappaB nuclear translocation and there is a possibility that this prevention of p65 nuclear translocation may involve p38 and ERK MAPKs as cross talks occur between MAPK and NF-kappaB pathways. This study provides an insight into the probable mechanism(s) underlying the anti-inflammatory and therapeutic properties of betulinic acid.

Molecular mechanisms of anti-inflammatory action of the flavonoid, tricin from Njavara rice (Oryza sativa L.) in human peripheral blood mononuclear cells: possible role in the inflammatory signaling.

Flavonoids are a group of natural substances that are located in sources of vegetal origin and are able to regulate acute and chronic inflammatory responses. The anti-oxidant and anti-inflammatory effects corroborate with the preferential use of Njavara, a rice variety in indigenous medicine and the phytochemical investigations revealed the occurrence of a flavonoid, tricin at significantly higher levels compared to staple varieties. This study describes the new aspects of inflammatory suppression by the Njavara rice by evaluating the role of active constituent, tricin in the regulation of production of various pro-inflammatory markers by human peripheral blood mononuclear cells stimulated with lipopolysaccharide. Treatment with tricin resulted in significant down-regulation of LPS-elicited production of TNF-α, IL-6, PGE(2) and NO. Tricin was found to be a potential blocker of the expression of isoforms of nitric oxide synthase, cyclooxygenase and matrix metalloproteinases. Modulation of the cascade of molecular events in lipopolysaccharide signaling also includes inhibition of transcription factor NF-κB evidenced by the detection of enhanced p65 subunit in the nuclear extracts on tricin supplementation. The present study summarizes the role of the flavonoid, tricin in the modulation of the expression of different inflammatory mediators and revealed that the inhibitory effects on cell signaling pathways are responsible for its anti-inflammatory activity.

Inhibitory effect of Ruta graveolens L. on oxidative damage, inflammation and aortic pathology in hypercholesteromic rats.

The purpose of the study was to investigate the efficacy of methanolic extract of Ruta graveolens L. in reducing oxidative damage, inflammation and aortic pathology in hypercholesteremic rats. For the study rats were divided into three groups - control group, hypercholesteremic group and treatment group (20 mg MER/kg/d orally) - and were fed for 90 days. Serum total cholesterol, LDL-C, total WBC count, CRP level, TBARS, atherogenic index, activities of COX, 15 LOX in monocyte and serum myeloperoxidase were increased in cholesterol fed rats. Activities of antioxidant enzymes and the concentration reduced glutathione in liver and heart tissue and serum HDL-C were decreased in cholesterol fed rats. The results showed that level of total cholesterol, LDL-C, atherogenic index was decreased and HDL-C was increased in MER treated rats. Activities of antioxidant enzymes were found to be increased and the activity of MPO, COX and 15 LOX were decreased on supplementation with MER. Concentration of TBARS and total WBC count were decreased and GSH was increased on supplementation with MER. Histopathology of aorta of cholesterol fed rat showed marked alterations whereas the aorta of MER administrated rat showed no significant changes. These results suggested that MER reduces oxidative stress, inflammation and aortic pathology in hypercholesteremic rats. Thus the plant may therefore be useful for therapeutic treatment of clinical conditions associated atherosclerosis.

15(S)-HETE-induced angiogenesis in adipose tissue is mediated through activation of PI3K/Akt/mTOR signaling pathway

Chronic low-grade inflammation underlies obesity and associated metabolic dysfunctions. Lipoxygenase pathways are activated in adipose tissue during obese conditions. Since adipogenesis is associated with angiogenesis, the present study was designed to examine the role of 15-lipoxygenase metabolite, 15(S)-hydroxyeicosatetraenoic acid [15(S)-HETE] on angiogenesis in adipose tissue. Results showed that 15(S)-HETE induced sprouting in fat pad stromovascular tissues, induced morphological changes relevant to angiogenesis in endothelial cells derived from adipose tissue, upregulated the production of CD31, upregulated the gene level expression and production of vascular endothelial growth factor (VEGF), indicating the pro-angiogenic effect of 15(S)-HETE. LY294002, an inhibitor of PI3K-Akt pathway, and rapamycin, inhibitor of mammalian target of rapamycin (mTOR), significantly reversed the effect of 15(S)-HETE. 15(S)-HETE also induced activation of Akt and mTOR. These observations suggest that 15(S)-HETE stimulates angiogenesis in adipose tissue through activation of PI3K/Akt/mTOR signaling.

Anti-inflammatory potential of an ethyl acetate fraction isolated from Justicia gendarussa roots through inhibition of iNOS and COX-2 expression via NF-κB pathway

Justicia gendarussa Burm.f. (J. gendarussa) is a plant used as traditional medicine in different parts of India and China to treat inflammatory disorders like rheumatoid arthritis. But its mechanism of anti-inflammatory action is still unclear. Hence in this context, the objective of our study is to reveal the mechanism of anti-inflammatory activity of J. gendarussa which would form an additional proof to the traditional knowledge of this plant. The anti-inflammatory function and mechanism(s) of action was studied in an ethyl acetate fraction isolated from methanolic extract of J. gendarussa roots (EJG). Anti-inflammatory studies were conducted on rats using partitioned fractions isolated from methanolic extract of J. gendarussa roots. In carrageenan-induced rat paw edema, ethyl acetate fraction brought about 80% and 93% edema inhibition at 3rd and 5th hour at a dose of 50 mg/kg, when compared to other extracts and Voveran. We investigated whether EJG inhibits the release of cycloxygenase (COX), 5-lipoxygenase (5-LOX), interleukin-6 (IL-6) and nuclear factor kappa B (NF-κB) in LPS stimulated human peripheral blood mononuclear cells (hPBMCs). Results shows that EJG dose dependently inhibited LPS-activated COX, 5-LOX, IL-6, and NF-κB in hPBMCs. EJG also reduced LPS induced levels of iNOS and COX-2 mRNA expression in hPBMCs. This study provides an insight into the probable mechanism(s) underlying the anti-inflammatory activity of EJG and therefore, we report the first confirmation of the anti-inflammatory potential of this traditionally employed herbal medicine in vitro.

Inhibition of pro-inflammatory mediators: role of Bacopa monniera (L.) Wettst

Bacopa monniera (L.) Wettst is a renowned plant in the Ayurvedic system of medicine. The present study seeks to identify the anti-inflammatory activity of two fractions from the methanolic extract of Bacopa, viz. the triterpenoid and bacoside-enriched fractions. The ability of these two fractions to inhibit the production of pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-6 was tested using lipopolysaccharide (LPS)-activated peripheral blood mononuclear cells and peritoneal exudate cells in vitro. We found that triterpenoid and bacoside-enriched fractions significantly inhibited LPS-activated TNF-α, IL-6 and nitrite production in mononuclear cells. Significant antioxidant activity was exhibited by the bacoside enriched fraction compared to the triterpenoid fraction. Carrageenan-induced hind paw oedema assay revealed that triterpenoid and bacoside-enriched fractions exerted anti-oedematogenic effect, while in the arthritis model only the triterpenoid fraction exerted an anti-arthritic potential. The present study provides an insight into the ability of Bacopa monniera to inhibit inflammation through modulation of pro-inflammatory mediator release.

Inhibitory effects of Cynodon dactylon L. on inflammation and oxidative stress in adjuvant treated rats

Cynodon dactylon is one of the 10 auspicious herbs that constitute the group Dasapushpam in Ayurveda. Traditionally Cynodon dactylon L. is used against many chronic inflammatory diseases in India. The present study was carried out to evaluate the protective effect of Cynodon dactylon against rats with adjuvant- induced arthritis. Arthritis was induced by intradermal injection of complete Freund’s adjuvant into the right hind paw produce inflammation of the joint. A significant increase in the levels of inflammatory mediators, myeloperoxidase, nitrite, C-reactive protein, ceruloplasmin was observed. This was associated with oxidative stress with a marked reduction in the activity of catalase, superoxide dismutase, glutathione peroxidase and the levels of glutathione, vitamins C and E and an increase in the lipid peroxidation as indicated by the higher levels of thiobarbituric acid reactive substances. Cynodon dactylon (20mg/kg/b.wt) was orally administered to arthritic rats after adjuvant injection produced a significant attenuation in the inflammatory response, oxidative stress and ameliorated the arthritic changes to near normal conditions. Hence, the results of this study clearly indicate that Cynodon dactylon extract has a promising protective role against arthritis.