![]() (E) Analysis of total COX-enzymatic activity (COX1/2) comparing cells either left untreated (control, UT) or treated with TAK inhibitor (5 μM) (control, black bars) with cells incubated with different concentrations (as indicated) of comfrey-OP, diclofenac, or the COX-1 inhibitor SC-560. Total COX enzymatic activity is depicted compared to untreated cells. ![]() In vitro analysis of recombinant COX-1 enzymatic activity comparing samples which were either left untreated (control, UT) or treated with the specific COX-1 inhibitor SC-560 (control, black bars) with samples incubated with different concentrations of comfrey-OP (gray bars) or diclofenac (white bars). (D) Comfrey-OP does not affect COX-1 activity. Expression levels are depicted as mean fold change compared to non-stimulated cells. Relative mRNA levels of COX-2 were normalized to GAPDH. (C) Real-time PCR of IL-1β induced (5 ng/ml time points of stimulation as indicated) HUVEC either left untreated (CTRL, squares) or pretreated (30 min) with comfrey-RE (200 μg/ml boxes) and comfrey-OP (20 μg/ml asterisks). Treatment of cells with comfrey preparations (RE and OP) reduces COX-2 protein levels. (B) Western blot analysis of samples used in A showing that COX-2 expression can only be found in IL-1β induced samples. Note that both comfrey preparations diminish the IL-1β induced COX-2 activity (black bars), but not the constitutive COX-1 activity (gray bars). Total COX enzymatic activity is depicted compared to non-stimulated, untreated cells. (A) Analysis of total COX-enzymatic activity (COX1/2) comparing unstimulated (UT, gray bars) and IL-1β (5 ng/ml, 90 min) stimulated HUVEC (black bars) which were either left untreated (control) or pretreated (30 min) with comfrey-RE (200 μg/ml), comfrey-OP (20 μg/ml), or TAK inhibitor (5 μM). Relative mRNA levels of E-selectin were normalized to GAPDH and values are depicted as mean fold change compared to non-stimulated cells. ![]() (C) Real-time PCR of IL-1β induced (5 ng/ml time points of stimulation as indicated) HUVEC either left untreated (CTRL, circles) or pretreated (30 min) with comfrey-RE (200 μg/ml squares), comfrey-OP (20 μg/ml asterisks) or the TAK1 inhibitor (5Z)-7-oxozeaenol (used as positive control at 5 μM). E-selectin protein levels were analyzed after 2 h. (B) Cell ELISA of IL-1β (5 ng/ml, 90 min) stimulated HUVECs which were either left untreated (CTRL, black bar) or pretreated (30 min) with different doses (either 160, 130, 80, 60 μg/ml) of comfrey-RE (gray bars), comfrey-OP (20, 12, 10, 6 μg/ml dark gray bars) or the inhibitor PHA-408 (20, 10, 5, 2.5 μM light gray bars). Relative mRNA levels of E-selectin were normalized to GAPDH and expression levels are depicted as mean fold change compared to non-stimulated cells. (A) Real-time PCR of IL-1β (5 ng/ml, 90 min) stimulated HUVEC which were either left untreated (CTRL, black bar) or pretreated (30 min) with either 166, 125, 100 μg/ml of comfrey-RE (gray dotted bars) or with 20, 10, 5 μg/ml of comfrey-OP (black dotted bars). officinale root extract preparations affect E-selectin expression in a dose- and time-dependent manner. NF-κB Symphytum officinale comfrey endothelial cells inflammation transactivation transcription. These results provide a first mechanistic insight into the mode of action of a century-old popular herbal medicine. Furthermore, our biochemical studies provide evidence that comfrey inhibits NF-κB signaling at two stages: it dampens not only the activation of IKK1/2 and the subsequent IκBα degradation, but also interferes with NF-κB p65 nucleo-cytoplasmatic shuttling and transactivation. Both preparations inhibit the activation of NF-κB, a transcription factor of central importance for the expression of these and other pro-inflammatory genes. The extract, and especially its mucilage-depleted fraction, impair the interleukin-1 (IL-1) induced expression of pro-inflammatory markers including E-selectin, VCAM1, ICAM1, and COX-2. Here, we show that a hydroalcoholic extract of comfrey root impairs the development of a pro-inflammatory scenario in primary human endothelial cells in a dose-dependent manner. However, the molecular basis of its action remained elusive. Today, its topical use is based on its analgesic and anti-inflammatory effects, which have been substantiated by modern clinical trials. Symphytum officinale, commonly known as comfrey, constitutes a traditional medicinal plant with a long-standing therapeutic history, and preparations thereof have been widely used for the treatment of painful muscle and joint complaints, wound and bone healing, and inflammation.
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