大蒜萃取物對小鼠巨噬細胞株一氧化氮、細胞激素和前列腺素生成的
影響
Effects of garlic extracts on the production of nitric oxide, cytokines and prostaglandin in RAW 264.7 cells
中文摘要
本研究主要以小鼠巨噬細胞RAW 264.7 為實驗模式，探討大蒜萃取物 diallyl sulfide（DAS）、diallyl disulfide（DADS）與 allyl methyl sulfide（AMS）對細菌 內毒素lipopolysaccharide（LPS）所誘導之發炎相關因子一氧化氮（nitric oxide, NO）、腫瘤壞死因子（tumor necrosis factor-α, TNF-α）、白細胞介素 1β
（interleukin-1β, IL-1β）、interleukin-6（IL-6）、interleukin-10（IL-10）、前列腺 素（prostaglandin E2, PGE2）生成的影響，並進一步瞭解其可能機制。結果顯示，
DAS（0.02?10 μM）、DADS（0.01?0.5 μM）與 AMS（0.2?20 μM）對細胞 皆不會與AMS（2?20 μM）可抑制由 LPS 所誘導之 NO 的生成並呈現劑量效應，
其中以DADS 之作用最為明顯。對細胞激素的結果顯示，較高濃度之 DAS 可抑
制由LPS 所誘導之 TNF-α、IL-1β、IL-6、IL-10 的生成；DADS 可抑制由 LPS 所誘導之TNF-α、IL-10 的生成；AMS 可抑制由 LPS 所誘導之 IL-1β的生成以 及促進IL-10 的生成。除此之外 DAS 可抑制由 LPS 所誘導之 PGE2 的生成，DADS 和AMS 則對 LPS 所誘導之 PGE2 的生成無影響。Western blot 分析結果指出，
DAS、DADS 可抑制細胞中 iNOS (inducible nitric oxide synthase)蛋白質的表現並 呈現劑量關係，且高濃度之DAS 亦可抑制細胞中 COX-2 (cyclooxgenase-2)蛋白
質的表現，這與其抑制NO 與 PGE2 的生成有一相對應關係。除此之外，DAS、
DADS、AMS 具直接清除 NO 的作用。綜合以上結果得知，大蒜萃取物 DAS、
DADS 與 AMS 對 LPS 所誘導之 NO、PGE2 及細胞激素的分泌有不同之作用，
其中DAS 可部分藉由抑制 COX-2、iNOS 蛋白質的表現而降低由 LPS 誘導之 PGE2、NO 的生成，且同時伴隨抑制由 LPS 所誘導之發炎因子 TNF-α、IL-1β、
IL-6 與 IL-10 的生成；DADS 可部分藉由抑制 iNOS 蛋白質的表現而抑制由 LPS 誘導之NO 的生成，且同時伴隨抑制因 LPS 所誘導之發炎因子 TNF-α及 IL-10 的生成；而AMS 僅可降低由 LPS 誘導之 NO、TNF-α和 IL-1β的生成及促進 IL-10
的生成。然而，大蒜萃取物本身DAS、DADS 與 AMS 皆可藉由直接清除 NO 之
能力，而降低培養基中NO 的生成。
英文摘要
Garlic extracts, including diallyl sulfide (DAS), diallyl disulfide (DADS) and allyl methylsulfide (AMS), are thought to be the bioactive components in garlic. A lot of studies have demonstrated that garlic are effective food of anticancer, and its bioactive
components inhibit inflamation. In the present study, we investigated the garlic
extracts DAS, DADS and AMS on the inhibition of the nitric oxide, cytokines (TNF-α, IL-1β, IL-6, IL-10) and prostaglandins in lipopolysaccharide (LPS) stimulated RAW 264.7 cells. The results showed that DAS (0.02?10 μM), DADS (0.01?0.5 μM) or AMS (0.2?20 μM) didn’t effect viability of RAW 264.7 cells. The results showed DAS (0.1?10 μM), DADS (0.05?0.5 μM) or AMS (2?20 μM) had a significant dose-dependent inhibition on nitric oxide production in RAW 264.7 cells. DAS had better inhibition than DADS and AMS. Parallel to the NO production, cytokines analysis indicated that DAS (1?10 μM) could inhibit LPS induced the produciotn of TNF-α, IL-1β, IL-6 and IL-10 in a dose-depentdent manner (p<0.05). DADS could inhibit the produciotn of TNF-α and IL-10. AMS could inhibit IL-1β produciotn and induce IL-10 produciotn. Another DAS could inhibit LPS induced PGE2 produciotn, and DADS and AMS had no significantly inhibitory effect. Western blot analysis indicated that high concentration of DAS and DADS could inhibit LPS induced expression of inducible nitric oxide (iNOS) protein. DAS could slightly inhibit expression of cyclooxygenase-2 (COX-2) protein. However, DADS and AMS didn’t have significantly inhibitory effect. This results could parallel to inhibit the
production of NO and PGE2. Besides DAS, DADS and AMS itself could inhibit the production of NO in medium. In conclusion, garlic extracts, DAS, DADS and AMS had differential inhibitory patterns on the production of NO, cytokins and PGE2. DAS inhibited LPS induced production of NO, PGE2, TNF-α, IL-1β, IL-6 and IL-10.
DADS, to a lesser extent, inhibited LPS induced produciton of NO, TNF-α and IL-10.
However, AMS only inhibited the production of NO, TNF-α and IL-1β, but induced IL-10 production. Besides, DAS, DADS and AMS directly lowered NO content in culture medium by their scavenging ability.
