Abstract
Inflammatory pain is often difficult to treat, and consequently there is a need to develop novel analgesic drugs that target novel or known targets. Studies carried out in-vitro suggest that Gingko biloba may have anti-inflammatory properties and modulatory effects on key pain-related signalling molecules, and therefore may be a potential novel analgesic drug. The aim of this project was to investigate the therapeutic potential in-vivo and molecular mechanisms in-vitro of EGb-761, a standardised extract of Ginkgo biloba, as a novel agent in the treatment of inflammatory pain. To further elucidate the mechanisms of action of EGb-761, the anti-inflammatory properties of the individual constituents (quercetin, kaempferol, isorhamnetin, ginkgolide A, B and C and bilobalide) were also investigated.The analgesic properties of EGb-761 were investigated in two well characterised models of inflammatory pain, hyperalgesia and allodynia; intradermal injection of carrageenan into the hindpaw and hindpaw incision. The effects of EGb-761 administered orally (3 - 300 mg kg'1; p.o.), intraplantarly (100 and 300 pg; i.pl.) or intrathecally (0.5 and 1 pg; i.t.) 30 minutes pre- or 3 or 21 hours post-injury on hindpaw withdrawal latency (in secs) to thermal stimulation, response threshold (in grams) to mechanical stimulation and paw oedema (cm3) were measured in adult male rats before and up to 7 days post-carrageenan/incision. EGb-761 administered orally dose-dependently inhibited carrageenan- and hindpaw incision induced thermal hyperalgesia but not mechanical allodynia or paw oedema in both pain models, and the effectiveness was dependent on time of administration; the antihyperalgesic effect being more pronounced when EGb-761 was administered 3 or 21 hours post-injury. Intrathecal and intraplantar injection of EGb-761 also dose-dependently inhibited carrageenan-induced thermal hyperalgesia and paw oedema but not mechanical allodynia.
The molecular mechanisms of EGb-761 and the individual constituents’ anti-inflammatory effects were tested in-vitro in a model of lipopolysaccharide (LPS) induced inflammation in murine RAW264.7 macrophages. The effects of treatment with EGb-761, or the individual constituents, quercetin, kaempferol and isorhamnetin and ginkgolide A, B and C and bilobalide, or drug-vehicle for 24 h on LPS-induced up-regulation of key inflammatory pain signalling molecules tumor necrosis factor alpha (TNF-a), cyclooxygenase 1 and 2 (COX-1, COX-2), inducible nitric oxide synthase (iNOS) and nitric oxide (NO) were measured using real-time PCR, Western blotting and the Griess reaction. EGb-761 reduced LPS-induced TNF-a mRNA, COX-2 and iNOS mRNA and protein and NO release but not COX1 mRNA. Each of the constituents, with the exception of ginkgolide C, inhibited COX-2 mRNA induction to a similar extent as EGb-761. Kaempferol, isorhametin, ginkgolide B and bilobalide all decreased iNOS mRNA to a similar extent as EGb761 but only kaempferol and isorhamnetin decreased NO release while quercetin increased NO release. The constituents failed to inhibit induction of TNF-a mRNA, while quercetin increased TNF-a mRNA expression. These data indicate that multiple constituents (but not ginkgolide C), contribute to the EGb-761’s anti-inflammatory effect mainly through targeted inhibition of the COX-2 and the NO signalling pathway.
Together these results demonstrate that EGb-761 has significant analgesic and anti-inflammatory properties and that this therapeutic action is mediated via a peripheral and central site of action. Furthermore, evidence suggests that EGb-761 may mediate these anti-inflammatory and analgesic effects via inhibition of COX-2 and NO signalling pathways at central and peripheral sites.
| Date of Award | 2008 |
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| Original language | English |
| Awarding Institution |
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| Supervisor | Sharron Dolan (Supervisor) |