Synergistic protective effect of curcumin and resveratrol against oxidative stress in endothelial EAhy926 cells

Curcumin (C) and resveratrol (R) are two well-known nutraceuticals with strong antioxidant activity that can protect cells from oxidative stress. This study aims to investigate the synergy of CR combinations in protecting human endothelial EAhy926 cells against H2O2-induced oxidative stress and its related mechanisms. C and R as individual compounds as well as CR combinations at different ratios were screened for their protective effects against H2O2 (2.5 mM) induced cell death assessed by cell viability assays. The synergistic interaction was analysed using the combination index model. The effects of optimal CR combinations on caspase-3 activity, ROS level, SOD activity, NAD cellular production, expression of Nrf2 and HO-1, and Nrf2 translocation were determined. CR combinations produced a synergistic protection against that of H2O2-induced changes in cell viability, caspase-3 activity, and ROS production. The strongest effect was observed for CR with the ratio of 8 : 2. Further experiments showed that CR 8 : 2 exhibited significantly greater effects in increasing Nrf2 translocation and expressions of Nrf2 and HO-1 proteins, as well as SOD activity and total cellular NAD production, than that of C or R alone. The findings demonstrate that combination of C and R produced a strong synergy in activity against H2O2-induced oxidative stress in EAhy926 cells. The mechanism of this synergy involves the activation of Nrf2-HO-1 signaling pathway and promotion of antioxidant enzymes. Further studies on CR synergy may help develop a new combination therapy for endothelial dysfunction and other conditions related to oxidative stress

Synergistic anti-inflammatory activity of ginger and turmeric extracts in inhibiting lipopolysaccharide and interferon-γ-induced proinflammatory mediators

This study aims to investigate the combined anti-inflammatory activity of ginger and turmeric extracts. By comparing the activities of individual and combined extracts in lipopolysaccharide and interferon-gamma-induced murine RAW 264.7 cells, we demonstrated that ginger-turmeric combination was optimal at a specific ratio (5:2, w/w) in inhibiting nitric oxide, tumour necrosis factor and interleukin 6 with synergistic interaction (combination index < 1). The synergistic inhibitory effect on TNF was confirmed in human monocyte THP-1 cells. Ginger-turmeric combination (5:2, w/w) also upregulated nuclear factor erythroid 2-related factor 2 activity and heme oxygenase-1 protein expression. Additionally, 6-shogaol, 8-shogaol, 10-shogaol and curcumin were the leading compounds in reducing major proinflammatory mediators and cytokines, and a simplified compound combination of 6-s, 10-s and curcumin showed the greatest potency in reducing LPS-induced NO production. Our study provides scientific evidence in support of the combined use of ginger and turmeric to alleviate inflammatory processes

Synergistic inhibition of pro-inflammatory pathways by ginger and turmeric extracts in RAW 264.7 cells

Synergy plays a prominent role in herbal medicines to increase potency and widen the therapeutic windows. The mechanism of synergy in herbal medicines is often associated with multi-targeted behavior and complex signaling pathways which are challenging to elucidate. This study aims to investigate the synergistic mechanism of a combination (GT) of ginger (G) and turmeric (T) extracts by exploring the modulatory activity in lipopolysaccharides (LPS)-induced inflammatory pathways and key molecular targets. A Bioplex ProTM mouse cytokine 23-plex assay was utilized to assess the broad anti-cytokine activity of GT in LPS and interferon (IFN)-gamma (both at 50 ng/mL)-activated RAW 264.7 cells. The inhibitory effects of individual and combined G and T on major proinflammatory mediators including nitric oxide (NO), tumor necrosis factor (TNF) and interleukin (IL)-6 were tested using Griess reagents and ELISA assays, respectively. Immunofluorescent staining and Western blot were used to investigate the modulatory effect of GT on key proteins in the LPS/TLR4 signaling transduction. The regulation of murine microRNA miR-155-5p was tested using real-time PCR. The IC50 value and combination index (CI) values were used to demonstrate potency and synergistic interaction, respectively. GT synergistically attenuated a range of pro-inflammatory mediators including inducible NO, major cytokines (TNF and IL-6) and secondary inflammatory cytokines (GM-CSF and MCP-1). GT significantly inhibited LPS-induced NF-kB p65 translocation, the activation of TLR4, TRAF6, and phosphorylation of JNK and c-JUN. Moreover, the suppressive effect of GT on each of the protein targets in this axis was stronger than that of the individual components. Real-time PCR analysis showed that GT suppressed miR-155-5p to a greater extent than G or T alone in LPS-stimulated cells. Our study demonstrates the synergistic mechanism of GT in downregulating LPS-induced proinflammatory pathways at the miRNA and protein levels. Our results establish a scientific basis for the combined application of G and T as an advanced therapeutic candidate in inflammatory diseases with broad and synergistic anti-inflammatory activity and multi-targeted mechanisms

Protective effect of the curcumin-baicalein combination against macrovascular changes in diabetic angiopathy

Endothelial dysfunction is an early pathological event in diabetic angiopathy which is the most common complication of diabetes. This study aims to investigate individual and combined actions of Curcumin (Cur) and Baicalein (Bai) in protecting vascular function. The cellular protective effects of Cur, Bai and Cur+Bai (1:1, w/w) were tested in H2O2 (2.5 mM) impaired EA. hy926 cells. Wistar rats were treated with vehicle control as the control group, Goto-Kakizaki rats (n=5 each group) were treated with vehicle control (model group), Cur (150 mg/kg), Bai (150 mg/kg), or Cur+Bai (75 mg/kg Cur + 75 mg/kg Bai, OG) for 4 weeks after a four-week high-fat diet to investigate the changes on blood vessel against diabetic angiopathy. Our results showed that Cur+Bai synergistically restored the endothelial cell survival and exhibited greater effects on lowering the fasting blood glucose and blood lipids in rats comparing to individual compounds. Cur+Bai repaired the blood vessel structure in the aortic arch and mid thoracic aorta. The network pharmacology analysis showed that Nrf2 and MAPK/JNK kinase were highly relevant to the multi-targeted action of Cur+Bai which has been confirmed in the in vitro and in vivo studies. In conclusion, Cur+Bai demonstrated an enhanced activity in attenuating endothelial dysfunction against oxidative damage and effectively protected vascular function in diabetic angiopathy rats

[In Press] Identification of Nrf2 activators from the roots of Valeriana officinalis

Various age-related chronic diseases have been linked to oxidative stress. The cellular antioxidant response pathway is regulated by the transcription factor Nrf2. Therefore, plant-derived Nrf2 activators might be useful therapeutics to stimulate the body's defense mechanisms. Our study was focused on the discovery of potent Nrf2 activators from medicinal plants. Initially, a variety of medicinal plant extracts were screened for Nrf2 activity using an Nrf2 luciferase reporter cell line. Among these, Valerian (V. officinalis) root was identified as a potent candidate. Sequential extraction and bioassay-guided fractionation led to the isolation of four Nrf2-active compounds, which were structurally identified by NMR and LC/HRMS as the known compounds isovaltrate, valtrate, jatamanvaltrate-P, and valerenic acid. These four compounds were then tested in relevant biological assays. Firstly, their effects on the expression of glutathione S-transferase (GST), glutamate—cysteine ligase catalytic subunit (GCLC), glutathione peroxidase (GPX), and heme oxygenase 1 (HO-1) were determined in HepG2 cells. GSTP1 and GCLC were upregulated by isovaltrate, valtrate and jatamanvaltrate-P, HO-1 by isovaltrate, jatamanvaltrate-P and valerenic acid. The four compounds also increased the levels of glutathione (GSH) and its metabolite, CysGly. As GSH aids in the detoxification of H2O2, cytoprotective effects of these four Nrf2 activators against H2O2 toxicity were investigated, and indeed, the compounds significantly improved cell survival. This study provides evidence that four valepotriates from the roots of V. officinalis are activators of Nrf2-mediated antioxidant and detoxification pathways. Our data might expand the medical use of this plant beyond its current application as a sleep aid

Combined effect of 2 nutraceuticals, curcumin and resveratrol on the activation of nuclear factor erythroid 2-related factor 2

Purpose: Combination therapy presents a systematic approach in tackling complex pathological conditions such as cardiovascular diseases (CVD). Curcumin (a polyphenol found in Curcuma longa) and resveratrol (a stilbenoid found in strongly pigmented vegetables and fruit) are 2 popular nutraceuticals known for their powerful antioxidant activities. The nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor that responds to oxidative stress by binding to antioxidant response element (ARE). As isolated compounds, curcumin and resveratrol, are common Nrf2 activators; however, their combined effect has yet to be determined. Methods: AREc32 cells were culture and coincubated with individual or combined curcumin and resveratrol (0– 100 mM). After 24 h co-incubation, the cells were washed and replaced with luciferin buffer, and the expression of Nrf2 was analyzed using bioluminescence. Alamar blue assay was used to determine the cell viability. Results: Our results suggested that curcumin-resveratrol, at both 50 and 25 mM, significantly upregulated Nrf2 expression compared to negative control (cells with medium only) (P <.05), without causing any cytotoxicity. Conclusion: The combined activity of curcumin and resveratrol against oxidative stress provides early evidence of a combination therapy that will assist in the management of various chronic conditions such as CVD

6-shogaol and 10-shogaol synergize curcumin in ameliorating proinflammatory mediators via the modulation of TLR4/TRAF6/MAPK and NFkB translocation

Extensive research supported the therapeutic potential of curcumin, a naturally occurring compound, as a promising cytokinesuppressive anti-inflammatory drug. This study aimed to investigate the synergistic anti-inflammatory and anti-cytokine activities by combining 6-shogaol and 10-shogaol to curcumin, and associated mechanisms in modulating lipopolysaccharides and interferon-ɣ-induced proinflammatory signaling pathways. Our results showed that the combination of 6-shogaol-10-shogaolcurcumin synergistically reduced the production of nitric oxide, inducible nitric oxide synthase, tumor necrosis factor and interlukin-6 in lipopolysaccharides and interferon-γ-induced RAW 264.7 and THP-1 cells assessed by the combination index model. 6-shogaol-10-shogaol-curcumin also showed greater inhibition of cytokine profiling compared to that of 6-shogaol-10-shogaol or curcumin alone. The synergistic anti-inflammatory activity was associated with supressed NFκB translocation and downregulated TLR4-TRAF6-MAPK signaling pathway. In addition, SC also inhibited microRNA-155 expression which may be relevant to the inhibited NFκB translocation. Although 6-shogaol-10-shogaol-curcumin synergistically increased Nrf2 activity, the anti-inflammatory mechanism appeared to be independent from the induction of Nrf2. 6-shogaol-10-shogaol-curcumin provides a more potent therapeutic agent than curcumin alone in synergistically inhibiting lipopolysaccharides and interferon-γ induced proinflammatory mediators and cytokine array in macrophages. The action was mediated by the downregulation of TLR4/TRAF6/MAPK pathway and NFκB translocation