Ginkgo biloba extract enhances antiplatelet and antithrombotic effects of cilostazol without prolongation of bleeding time.

Thrombosis and thromboembolic occlusions of major and minor blood vessels are a major complication in various peripheral vascular diseases. Antiplatelet agents (APA), key tools in the treatment of atherothrombosis, therefore became a mainstay medication for a wide range of vascular diseases. Cilostazol and Ginkgo biloba extract (GB), commonly used remedies for peripheral arterial disease, inhibit platelet aggregation with distinct therapeutic mechanisms. In this study, we have investigated if GB can potentiate the antiplatelet effects of cilostazol to explore the utility of combination therapy of cilostazol and GB against peripheral occlusive vascular diseases. GB or cilostazol was evaluated alone or in combination for the antiplatelet activity using in vitro and in vivo models. In addition, potential bleeding side effect of the combinative therapy was assessed by measuring bleeding time, prothrombin time (PT) and activated partial thromboplastin time (aPTT) in vivo after oral administration. In in vitro assays using freshly isolated human platelets, the combination of cilostazol and GB showed superior inhibition of both the shear and the collagen-induced platelet aggregation to those of each drug alone. In accordance with these enhanced in vitro antiplatelet activities, the combinative therapy showed enhanced anti-thrombotic effects in in vivo pulmonary embolism model and arterial thrombosis model. In particular, the increase of survival rate in pulmonary embolism model by combination treatment of cilostazol (25 mg/kg) and GB (20 mg/kg) was higher more than two-fold of those of the respective drugs. Notably, the combination of cilostazol and GB did not show a significant effect on the bleeding time, PT and aPTT increase, suggesting that GB may potentiate the antiplatelet effect of cilostazol without the prolongation of bleeding time or coagulation time. With these studies, we suggest that combinative therapy of GB and cilostazol might offer enhanced anti-thrombotic efficacies without increasing side-effects.

SK-126, a synthetic compound, regulates the production of inflammatory cytokines induced by LPS in antigen-presenting cells.

A variety of mediators released by immune cells triggers or enhances specific aspects of the inflammatory response. Dendritic cells (DCs) play an essential role in the innate immune system by shaping the adaptive immune responses and by controlling the production of cytokines in response to inflammatory stimuli. In the present study, we investigated whether SK-126, a pyridine derivative based on gentianine originated from a natural product, can affect the LPS-induced inflammatory cytokine production in DC. Interestingly, treatment of mouse bone marrow-derived dendritic cells (BMDCs) and the murine dendritic cell line, DC 2.4, with SK-126 completely suppressed LPS-induced TNF-alpha expression at both transcriptional and protein levels. In contrast to TNF-alpha, SK-126 enhanced IL-10 expression at both transcriptional and protein levels. To determine signaling pathways involved in the regulation of inflammatory cytokines, we examined the involvement of MAPK and the transcription factor, NF-kappaB. SK-126 enhanced ERK1/2 and p38 activation following LPS stimulation, but it did not induce phosphorylation of SAPK/JNK and NF-kappaB. Also, STAT3 phosphorylation after LPS stimulation was increased by SK-126 to a large extent. Using specific inhibitors, we confirmed that SK-126 has dual effects in which it suppresses TNF-alpha production and enhances IL-10 production via the up-regulation of ERK1/2 and p38. Finally, LPS-induced inflammatory responses such as TNF-alpha production in vivo were significantly reduced by treatment with SK-126. Therefore, our findings suggest that SK-126 may be a useful drug candidate to treat inflammatory diseases in which pro- or anti-inflammatory cytokines play a significant role in their pathogenesis.

Effects of Brx-019 (acetic acid 3,6a,9-triacetoxy-6,6a,7, 11b-tetrahydro-indeno [2,1-c] chromen-10-yl ester), a Brazilin derivative, on T cell-mediated immune responses in multiple low dose streptozotocin-induced diabetic C57BL/6 male mice.

Brx-019 (acetic acid 3,6a,9-triacetoxy-6, 6a,7,11b-tetrahydro-indeno [2,1-c] chromen-10-yl ester) was derived from brazilin (CAS 474-07-7) during a trial designed to search for immunomodulators with lower toxicity and more effective immunomodulating activities than brazilin. Brx-019 was selected as a potential immunomodulator based on its effects on Concanavalin A (Con A)-induced proliferation of splenocytes and the 3-[14,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. Intraperitoneally administered Brx-019 significantly improved delayed type hypersensitivity and increased immunoglobulin M (IgM) plaque forming cells (PFCs) in multiple low dose streptozotocin-induced diabetic mice (MLDS-diabetic mice). This finding suggests that Brx-019 may increase suppressed humoral and cell-mediated immunity in type 1 diabetes. Brx-019 also significantly increased Con A- or alloantigen-induced proliferation of splenocytes, Con A-induced interleukin 2 (IL-2) production from splenocytes, and IL-2-induced proliferation of Con A-activated splenocytes in MLDS-diabetic mice. These results suggest that Brx-019 might improve immunity in diabetic mice by increasing IL-2 production in splenocytes and responsiveness of splenocytes to IL-2, which were suppressed in MLDS-diabetes.

Effects of gentianine on the production of pro-inflammatory cytokines in male Sprague-Dawley rats treated with lipopolysaccharide (LPS).

This study was undertaken to elucidate the mechanism of anti-inflammatory action of gentianine, a constituent of Gentiana Macrophylla. The effects of gentianine on lipopolysacharide (LPS)-induced production of pro-inflammatory cytokines were investigated in male Sprague-Dawley rats. For the first time, we found that oral administration of gentianine (10-100 mg/kg) suppressed the increases in tumor necrosis factor-alpha (TNF-alpha) (ED(50), 37.7 mg/kg) and interleukin (IL)-6 (ED(50), 38.5 mg/kg) in the sera from the rats challenged with bacterial LPS (100 microg/kg; i.p.). However, LPS induced production of other interleukins, such as IL-alpha, was not significantly altered by gentianine. These results suggest that the potential anti-inflammatory action of gentianine might be at least partly based on the suppressed production of TNF-alpha and IL-6.