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1 Buddlejasaponin IV isolated from Pleurospermum kamtschatidum is an anti-inflammatory compound that inhibits NO, PGE 2 and TNF-a production. Here, we studied the mode of action of this compound. 2 Buddlejasaponin IV (2.5-10 mM) reduced lipopolysaccaride (LPS (1 mg ml À1 ))-induced levels of iNOS and COX-2 at the protein levels, and iNOS, COX-2, TNF-a, interleukin (IL)-1b and IL-6 mRNA expression in RAW 264.7 macrophages in a concentration-dependent manner, as determined by Western blotting and RT-PCR, respectively. 3 Buddlejasaponin IV inhibited the LPS-induced activation of nuclear factor-kB (NF-kB), a transcription factor necessary for proinflammatory mediators, iNOS, COX-2, TNF-a, IL-1b and IL-6 expression. This effect was accompanied by a parallel reduction in IkB-a degradation and phosphorylation, and by the nuclear translocation of the NF-kB p65 subunit. 4 The effects of buddlejasaponin IV on acute phase inflammation were studied on serotonin-and carrageenan-induced paw edema. The antiedematous effect of buddlejasaponin IV was compared with 10 mg kg À1 of indomethacin p.o. Maximum inhibitions of 26 and 41% were noted at a dose of 20 mg kg À1 for serotonin-and carrageenan-induced paw edema, respectively. 5 The analgesic effect of buddlejasaponin IV was evaluated using acetic acid-induced writhing and hot-plate tests. Buddlejasaponin IV (10 and 20 mg kg À1 , p.o.) was found to have a marked analgesic effect in both models. 6 These results suggest that the inhibitions of the expressions of iNOS, COX-2, TNF-a, IL-1b and IL-6 by blocking NF-kB activation, are responsible for the anti-inflammatory effects of buddlejasaponin IV isolated from P. kamtschatidum.

We have investigated the anti-inflammatory effects of madecassic acid and madecassoside isolated from Centella asiatica (Umbelliferae) on lipopolysaccharide (LPS)-stimulated RAW 264.7 murine macrophage cells. Both madecassic acid and madecassoside inhibited the production of nitric oxide (NO), prostaglandin E(2) (PGE(2)), tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1beta), and IL-6. However, madecassic acid more potently suppressed these inflammatory mediators than did madecassoside. Consistent with these observations, madecassic acid inhibited the LPS-induced expression of iNOS and COX-2 at the protein level and of iNOS, COX-2, TNF-alpha, IL-1beta, and IL-6 at the mRNA level in RAW 264.7 macrophage cells, as determined by Western blotting and RT-PCR, respectively. Furthermore, madecassic acid suppressed the LPS-induced activation of nuclear factor-kappaB (NF-kappaB), and this was associated with the abrogation of inhibitory kappa B-alpha (IkappaB-alpha) degradation and with the subsequent blocking of p65 protein translocation to the nucleus. These results suggest that the anti-inflammatory properties of madecassic acid are caused by iNOS, COX-2, TNF-alpha, IL-1beta, and IL-6 inhibition via the downregulation of NF-kappaB activation in RAW 264.7 macrophage cells.

Nitric oxide (NO) which is produced by inducible nitric oxide synthase (iNOS) is a pro-inflammatory radical and an important biological mediator. 1) prostaglandin E 2 (PGE 2 ), released by arachidonic acid metabolites, is also an important mediator of acute and chronic inflammation, and it is associated with the enzyme, cyclooxygenase-2 (COX-2).2) In addition to these oxygen and arachidonic acid metabolites, tumor necrosis factor-a (TNF-a) acting as a cytokine or inflammatory mediator plays a major role in various inflammatory diseases, including septic shock and rheumatoid arthritis. 3,4) Nuclear factor-kB (NF-kB) appears to play a primary role in the transcriptional regulation of these iNOS, COX-2 and TNF-a gene expressions.5) NF-kB exists as a latent form in the cytoplasm of unstimulated cells and is bound to the inhibitory protein, IkB. 6) Phosphorylation of IkB leads to its degradation and the subsequent translocation of NF-kB to the nucleus where it activates transcriptions of target genes. 7,8) It is noteworthy that the use of medicinal plants or their crude extracts in the prevention and/or treatment of several chronic diseases has been traditionally practiced in different societies worldwide. Moreover, extracts of Ginkgo biloba LINNE (Ginkoaceae) have been used for centuries in traditional Chinese medicine. Today, a standardized extract (EGb 761) is prepared from Ginko biloba leaves and is prescribed commonly in Europe for the treatment of memory disorders, obstructive arteriosclerosis, Alzheimer's disease, ischaemic heart disease, cerebral infarction, aging, and age-related macular degeneration.9-11) Several modes of action of EGb 761 have been described: (1) its effects on the blood circulation, such as its vasoregulatory activity and rheological effects on blood (decreased viscosity, anti-platelet activating factor activity) 11) ; (2) its effects on metabolism changes, such as on neuron metabolism (increased tolerance to anoxia) 9,12,13) ; (3) its inhibition of cell membrane damage caused by free radicals 11,[13][14][15] ; and (4) its gene-regulatory effects, which suggest that it has anticancer activity. 16,17) EGb 761 contains two groups as its main active constituents: 24% flavonol glycoside (ginkgo-flavonol glycosides; quercetin, kaempferol, isorhamnetin) and 6% terpene (bilobalide and ginkgolides A, B, C).18) In the present study, we prepared a standardized Ginkgo biloba extract (GBB), i.e., an extract of Ginkgo biloba leaves standardized in terms of its total terpenes (12Ϯ3%), biflavonoid (4.5Ϯ1.5%), flavonol glycoside (Ͻ8%), and proanthocyanidine (under detection limit) content. The difference between GBB and EGb 761 is that it contains a higher level of terpene, a lower level of flavonol glycoside, and no detectable proanthocyanidine.In the present study, we compared the effects of these Ginkgo biloba extracts (GBB and EGb 761) on lipopolysaccharide (LPS)-induced NO and PGE 2 release. To further explore the possible mechanisms of these inhibitions by GBB, we investigated the expression levels...

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