Lignans From Forsythia x Intermedia Leaves and Flowers Attenuate the Pro-inflammatory Function of Leukocytes and Their Interaction With Endothelial Cells

Abstract

Aim of the study: Taking into account that overactivated leukocytes are an important factor in the development of many chronic diseases, we investigated the activity of phytochemically characterized (HPLC-DAD-MSn) extracts from forsythia leaves and flowers on the pro- and anti-inflammatory functions of leukocytes (effects on IL-1β, IL-8, TNF-α, and TGFβ release) and their adherence to endothelial cells. Using bio-guided fractionation, we isolated the active compounds and determined their biological activity, and we included the positive control quercetin.Methods: The effect on IL-1β, TNF-α, IL-8, and TGF-α production by leukocytes was measured by enzyme-linked immunosorbent assay (ELISA). The surface expression of adhesion molecules was analyzed with flow cytometry, and the neutrophil attachment to the endothelial cells was assessed fluorimetrically. The effects on p38MAPK, ERK1/2 and JNK phosphorylation were determined using western blots.Results: Leaf extracts had the effect of decreasing TNF-α production in neutrophils and monocyte/macrophage cells. The bio-guided fractionation led to the isolation of the following lignan aglycones: (+)-pinoresinol, (+)-epipinoresinol, (−)-matairesinol, (+)-phillygenin, and (−)-arctigenin. Only phillygenin was able to stimulate the anti-inflammatory function of macrophages by inducing TGF-β release and IL-10 receptor surface expression. Arctigenin, phillygenin, and a metabolite produced by the gut microbiota, enterolactone, decreased TNF-α and IL-1β production and neutrophil adhesion to endothelial cells, probably by attenuating the p38 and ERK kinase pathways.Conclusion: Forsythia x intermedia is a valuable source of active lignans, which may be potential candidates for treating inflammatory diseases that are associated with the excessive production of cytokines such as TNF-α and IL-1β.

UHPLC‐DAD‐ESI‐MS/MS and HPTLC profiling of ash leaf samples from different commercial and natural sources and their in vitro effects on mediators of inflammation

“…Compounds 46 ( t r = 32.2 min) and 52 ( t r = 34.2 min) exhibited pseudomolecular ions at m/z 519 [M‐H] − and m/z 565 [M‐H + HCOOH] − , respectively. According to the elution order in RP‐LC, compound 46 was tentatively identified as pinoresinol hexoside, while compound 52 was identified as epipinoresinol hexosides . Similar lignans were identified in the phloem of different Fraxinus sp …”

“…Fragmentation of this ion gave an ion at m/z 209 due to caffeoyl cleavage (loss of 162 amu), indicating that compounds 1 , 3 , 7 and 11 were caffeoyl hexaric acids. Hexaric acid has six diastereomers; however, in the Oleaceae family, only hydroxycinnamoyl‐glucaric and ‐altraric acids with different esterification patterns have been identified . Similarly, compounds 5 ( t r = 8.4 min), 8 ( t r =10.9 min), 10 ( t r = 11.7 min), 13 ( t r =13.6 min) and 18 ( t r =14.8 min) exhibited a pseudomolecular [M‐H] − ion at m/z 355.…”

“…Quercetin‐3‐ O ‐rutinoside was purchased from Roth (Karlsruhe, Germany). Oleuropein and verbascoside/acteoside were isolated as described previously . All solvents used for chromatography were of gradient grade.…”

“…UV spectra were recorded in the range 200–400 nm. Compounds were identified by comparing their retention time and UV‐visible and mass spectra with those obtained from standard compounds isolated previously from other Oleaceae family members and/or tentatively identified by comparison with literature information.…”

“…Peripheral venous blood was taken from healthy human donors (18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35) years old) at the Warsaw Blood Donation Centre. The donors did not smoke or take any medications and were clinically recognised to be healthy.…”