Anti-Inflammatory Activity of Bryophytes Extracts in LPS-Stimulated RAW264.7 Murine Macrophages.

Bryophytes produce rare and bioactive compounds with a broad range of therapeutic potential, and many species are reported in ethnomedicinal uses. However, only a few studies have investigated their potential as natural anti-inflammatory drug candidate compounds. The present study investigates the anti-inflammatory effects of thirty-two species of bryophytes, including mosses and liverworts, on Raw 264.7 murine macrophages stimulated with lipopolysaccharide (LPS) or recombinant human peroxiredoxin (hPrx1). The 70% ethanol extracts of bryophytes were screened for their potential to reduce the production of nitric oxide (NO), an important pro-inflammatory mediator. Among the analyzed extracts, two moss species significantly inhibited LPS-induced NO production without cytotoxic effects. The bioactive extracts of Dicranum majus and Thuidium delicatulum inhibited NO production in a concentration-dependent manner with IC50 values of 1.04 and 1.54 µg/mL, respectively. The crude 70% ethanol and ethyl acetate extracts were then partitioned with different solvents in increasing order of polarity (n-hexane, diethyl ether, chloroform, ethyl acetate, and n-butanol). The fractions were screened for their inhibitory effects on NO production stimulated with LPS at 1 ng/mL or 10 ng/mL. The NO production levels were significantly affected by the fractions of decreasing polarity such as n-hexane and diethyl ether ones. Therefore, the potential of these extracts to inhibit the LPS-induced NO pathway suggests their effective properties in attenuating inflammation and could represent a perspective for the development of innovative therapeutic agents.

Modulation of CD14 and TLR4·MD-2 activities by a synthetic lipid A mimetic.

Monosaccharide lipid A mimetics based on a glucosamine core linked to two fatty acid chains and bearing one or two phosphate groups have been synthesized. Compounds 1 and 2, each with one phosphate group, were practically inactive in inhibiting LPS-induced TLR4 signaling and cytokine production in HEK-blue cells and murine macrophages, but compound 3, with two phosphate groups, was found to be active in efficiently inhibiting TLR4 signal in both cell types. The direct interaction between compound 3 and the MD-2 coreceptor was investigated by NMR spectroscopy and molecular modeling/docking analysis. This compound also interacts directly with the CD14 receptor, stimulating its internalization by endocytosis. Experiments on macrophages show that the effect on CD14 reinforces the activity on MD-2·TLR4 because compound 3's activity is higher when CD14 is important for TLR4 signaling (i.e., at low LPS concentration). The dual targeting of MD-2 and CD14, accompanied by good solubility in water and lack of toxicity, suggests the use of monosaccharide 3 as a lead compound for the development of drugs directed against TLR4-related syndromes.