7,8-dihydroxyflavone displayed antioxidant effect through activating HO-1 expression and inhibiting caspase-3/PARP activation in RAW264.7 cells.

Flavonoids, which contain a benzo-γ-pyrone (C6-C3-C6) skeleton, have been reported to exhibit effective antioxidant ability. This study aimed to compare the antioxidant activities of 7,8-dihydroxyflavone (7,8-DHF) and 7-hydroxyflavone (7-HF) in H2 O2 , lipopolysaccharide (LPS), or tert-butyl hydroperoxide (t-BHP)-induced RAW264.7 cells, respectively. The antioxidant capacities of 7,8-DHF and 7-HF were firstly evaluated by 2,2-azinobis-3-ethyl-benzothiazoline-6-sulphonic acid (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) assays. Then, reactive oxygen species (ROS), super oxide dismutase (SOD), and malondialdehyde (MDA) productions in H2 O2 , LPS, or t-BHP-induced RAW264.7 cells were tested and compared, respectively. Finally, the antioxidant mechanisms of 7-HF and 7,8-DHF were initially investigated by western blot. Our results showed that 7,8-DHF possessed stronger free-radical scavenging capacity than 7-HF. Both 7,8-DHF and 7-HF suppressed MDA production and ROS accumulation, improved the activity of SOD in H2 O2 , LPS, or t-BHP-induced RAW264.7 cells, respectively. And 7,8-DHF exerted a better antioxidant effect than 7-HF, especially in t-BHP-induced oxidative stress. Mechanically, 7,8-DHF prevented the activation of poly ADP-ribosepolymerase and caspase-3, meanwhile markedly upregulated the expression of HO-1 protein in t-BHP-induced oxidative stress. These results suggested that 7,8-DHF might serve as a potential pharmaceutical drug against oxidative stress injury.

Diclofenac and eugenol hybrid with enhanced anti-inflammatory activity through activating HO-1 and inhibiting NF-κB pathway in vitro and in vivo.

A series of diclofenac hybrid molecules were synthesized and evaluated for their NO-inhibitory ability in LPS-induced RAW 264.7 macrophage cells. Among them, compound 1 showed the highest NO-inhibitory ability (approximately 66%) and no significant cytotoxicity. Compound 1 exhibited superior NF-κB-inhibitory ability compared to diclofenac through the activation of Nrf2/HO-1 signaling pathway in RAW 264.7. 20 mg/kg compound 1 resulted in remarkable colitis improvement in dextran sulfate sodium (DSS)-induced mice model by up-regulating HO-1 and down-regulating phosphorylation level of NF-κB p65. Moreover, 50 mg/kg dose of compound 1 showed a lower ulcerogenic potential compared to diclofenac in rats. The diclofenac-eugenol hybrid (compound 1) may serve as a novel anti-inflammatory agent based on its role in inhibiting the NF-κB signaling pathway and activating HO-1 expression with no toxicity in vitro and in vivo.

Design, synthesis, biological evaluation and molecular docking studies of novel pleuromutilin derivatives containing nitrogen heterocycle and alkylamine groups.

A series of pleuromutilin derivatives containing alkylamine and nitrogen heterocycle groups were designed and synthesised under mild conditions. The in vitro antibacterial activity of these semisynthetic derivatives against four strains of Staphylococcus aureus (MRSA ATCC 43300, S.aureus ATCC 29213, S.aureus AD3, and S.aureus 144) were evaluated by the broth dilution method. Compound 13 was found to have excellent antibacterial activity against MRSA (MIC = 0.0625 µg/mL). Furthermore, compound 13 was further studied by the time-killing kinetics and the post-antibiotic effect approach. In the mouse thigh infection model, compound 13 exhibited superior antibacterial efficacy than that of tiamulin. Meanwhile, compound 13 showed a lower inhibitory effect than that of tiamulin on RAW264.7 and 16HBE cells at the concentration of 10 µg/mL. Molecular docking study revealed that compound 13 can effectively bind to the active site of the 50S ribosome (the binding free energy = -9.66 kcal/mol).

Aesculetin exhibited anti-inflammatory activities through inhibiting NF-кB and MAPKs pathway in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Aesculetin (6,7-dihydroxy-2H-1-benzopyran-2-one) has been reported to exhibit potent anti-inflammatory property both in vitro and in vivo. AIMS OF THIS STUDY: In this study, we evaluated the anti-inflammatory effect and investigated underlying molecular mechanisms of aesculetin in LPS-induced RAW264.7 macrophages and DSS-induced colitis. MATERIALS AND METHODS: In this study, the production of NO, TNF-α, and IL-6 were measured to identify the aesculetin with potent anti-inflammatory effect. Then, the underlying anti-inflammatory mechanisms were explored by western blotting in LPS-induced cells. Next, we verify the anti-inflammatory potential of aesculetin in DSS-induced colitis in vivo. The clinical symptoms of colitis, including weight loss, DAI, colon length and MPO activity, and the secretion of TNF-α and IL-6 were evaluated. Finally, Western blot analysis was applied to further investigate underlying mechanism in DSS-induced colitis model. RESULTS: Our studies showed that aesculetin exhibited anti-inflammatory potential by inhibiting NO, TNF-α, and IL-6 production and reducing iNOS and NLRP3 expression in LPS-induced RAW264.7 cells. Mechanically, we found that aesculetin significantly inhibited LPS-induced activation of NF-κB and MAPKs signaling pathways. In DSS-induced mouse model, the colitis-related symptoms were relieved by treatment with aesculetin. Besides, aesculetin also inhibited the secretion of TNF-α and IL-6, and the activation of NF-κB and MAPKs signaling pathways in DSS-induced colitis. CONCLUSIONS: The anti-inflammatory effect of aesculetin was connected with its inhibition on the activation of NF-κB and MAPKs signaling pathways both in vitro and in vivo. Therefore, aesculetin was expected to be developed as an anti-inflammatory drug.

Design, synthesis and biological evaluation of novel pleuromutilin derivatives possessing 4-aminothiophenol linker as promising antibacterial agents.

A series of novel pleuromutilin derivatives containing 4-aminothiophenol moieties have been designed and synthesized as promising antibacterial agents against Methicillin-resistant Staphylococcus aureus (MRSA). The in vitro antibacterial activity of these semisynthetic derivatives against 4 strains of S. aureus (MRSA ATCC 43300, S. aureus ATCC 29213, S. aureus 144 and S. aureus AD3) was evaluated by the broth dilution method. Most of the synthesized derivatives displayed prominent in vitro activity (MIC ≤ 0.5 µg/mL). 12 Compounds possessed superior antibacterial activity against MRSA compared with valnemulin and retapamulin (MIC = 0.0625 µg/mL). Compounds 12, 16a, 16c and 19 exhibited the most effective antibacterial effect against MRSA (MIC = 0.015 µg/mL). Furthermore, the time-kill curves showed compounds 12 and 19 had a certain inhibitory effect against MRSA in vitro. Compounds 12 and 19 possessed longer PAE time (2.74 h and 3.11 h, respectively) than tiamulin (PAE = 2.04 h) against MRSA after exposure at 4 × MIC concentration for 2 h. Compounds 12 and 19 also displayed superior in vivo antibacterial efficacy (-1.20 log10 CFU/mL and -1.21 log10 CFU/mL, respectively) than tiamulin (-0.75 log10 CFU/mL) in reducing MRSA load in the mice thigh infection model. In addition, compound 19 had barely inhibitory effect on RAW 264.7 and 16HBE cells at 8 µg/mL. In molecular docking study, upon docking into the 50S ribosomal subunit, the binding free energy (ΔGb) of compound 12 and 19 was calculated to be -9.02 kcal/mol and -9.89 kcal/mol, respectively.

Discovery of Novel Pleuromutilin Derivatives as Potent Antibacterial Agents for the Treatment of MRSA Infection.

A series of novel pleuromutilin derivatives containing nitrogen groups on the side chain of C14 were synthesized under mild conditions. Most of the synthesized derivatives displayed potent antibacterial activities. Compound 9 was found to be the most active antibacterial derivative against MRSA (MIC = 0.06 µg/mL). Furthermore, the result of time-kill curves showed that compound 9 had a certain inhibitory effect against MRSA in vitro. Moreover, according to a surface plasmon resonance (SPR) study, compound 9 (KD = 1.77 × 10-8 M) showed stronger affinity to the 50S ribosome than tiamulin (KD = 2.50 × 10-8 M). The antibacterial activity of compound 9 was further evaluated in an MRSA-infected murine thigh model. Compared to the negative control group, tiamulin reduced MRSA load (~0.7 log10 CFU/mL), and compound 9 performed a treatment effect (~1.3 log10 CFU/mL). In addition, compound 9 was evaluated in CYP450 inhibition assay and showed only moderate in vitro CYP3A4 inhibition (IC50 = 2.92 µg/mL).

Design, synthesis, in vitro and in vivo evaluation against MRSA and molecular docking studies of novel pleuromutilin derivatives bearing 1, 3, 4-oxadiazole linker.

A class of pleuromutilin derivatives containing 1, 3, 4-oxadiazole were designed and synthesized as potential antibacterial agents against Methicillin-resistant staphylococcus aureus (MRSA). The ultrasound-assisted reaction was proposed as a green chemistry method to synthesize 1, 3, 4-oxadiazole derivatives (intermediates 85-110). Among these pleuromutilin derivatives, compound 133 was found to be the strongest antibacterial derivative against MRSA (MIC = 0.125 µg/mL). Furthermore, the result of the time-kill curves displayed that compound 133 could inhibit the growth of MRSA in vitro quickly (- 4.36 log10 CFU/mL reduction). Then, compound 133 (- 1.82 log10 CFU/mL) displayed superior in vivo antibacterial efficacy than tiamulin (- 0.82 log10 CFU/mL) in reducing MRSA load in mice thigh model. Besides, compound 133 exhibited low cytotoxicity to RAW 264.7 cells. Molecular docking studies revealed that compound 133 was successfully localized in the binding pocket of 50S ribosomal subunit (ΔGb = -10.50 kcal/mol). The results indicated that these pleuromutilin derivatives containing 1, 3, 4-oxadiazole might be further developed into novel antibiotics against MRSA.