探討DHMPC影響巨噬細胞及內皮細胞活化反應與趨化運動之機制

脂多醣體 (LPS, lipopolysaccharide) 是革蘭氏陰性菌外膜的主成分，能活化巨噬細胞及內皮細胞產生促發炎介質，趨化因子及黏附分子，進而調節巨噬細胞產生趨化運動。本論文使用小鼠巨噬細胞株 RAW264.7 及內皮細胞株 SVEC4-10 進行研究。文獻指出 DHMPC (5,7-dihydroxy-2-(4-methoxyphenyl) chromen-4-one) 能抑制發炎反應，至今並無研究顯示 DHMPC 對 RAW264.7 細胞的趨化運動有何影響，因此本論文探討 DHMPC 影響 LPS 誘導 RAW264.7 細胞及 SVEC4-10 細胞活化與其相關分子機制。結果發現，LPS 誘導 SVEC4-10 細胞活化，DHMPC 經由抑制 GEFs (guanine nucleotideexchange factors)，接著抑制 RhoGTPase，MAPKs (mitogen-activated protein kinases)，Akt 及 NF-κB (nuclear factor-κB) 路徑活化，減少促發炎介質及黏附分子產生，另一方面，抑制 RhoGTPase 活性進而抑制 LIMK 1/2 (Lin-11/Isl-1/Mec-3 kinase 1/2) 活化，使 cofilin 活化減少壓力絲產生，通透性下降及穩定接合點蛋白。此外，LPS 誘導 RAW264.7 細胞活化，DHMPC 經由抑制 GEFs 活性，增加 GAPs (GTPaseactivating proteins) 活性，抑制 RhoGTPase，ERK，Akt 及 NF-κB 路徑，減少產生促發炎介質及黏附分子，另一方面活化 LIMK 1/2 使 cofilin 去活化，抑制肌動蛋白延伸，因此減少 RAW264.7 細胞的黏著及遷移作用。本論文確定 DHMPC 抑制 RAW264.7 細胞及 SVEC4-10 細胞活化，而具有抗發炎效果。

Lipopolysaccharide (LPS), a major component of the outer membrane of Gram-negative bacteria, induces activation of macrophages and endothelial cells via expression of pro-inflammatory mediators, chemokines, adhesion molecules, and chemotaxis. We used the murine macrophage cell line, RAW264.7, and murine endothelial cell line SVEC4-10 for this study. Several studies have shown that DHMPC (5,7-dihydroxy-2-(4-methoxyphenyl)chromen-4-one) has anti-inflammatory effects. However, the effects of DHMPC in chemotaxis at RAW264.7 cells have never been demonstrated. The purpose of this study is to investigate the mechanisms of DHMPC in LPS-induced activation of RAW264.7 and SVEC4-10 cells. Our results show that DHMPC inhibits the activation of GEFs (guanine nucleotideexchange factors) in LPS-induced SVEC4-10 cells, and inhibits RhoGTPase, MAPKs (mitogen-activated protein kinases), Akt and NF-κB (nuclear factor-κB) activity. Furthermore, DHMPC down-regulates production of pro-inflammatory mediators and adhesion molecules. Also, DHMPC inactivates LIMK 1/2 (Lin-11/Isl-1/Mec-3 kinase 1/2) and activates cofilin resulting in degradation of stress fiber. Finally, DHMPC decreases permeability and maintains junctional complex of SVEC4-10 cells. In addition, DHMPC suppresss the activity of GEFs, augments GAPs (GTPaseactivating proteins) activation, and inactivates RhoGTPase, then inhibits ERK, Akt and NF-κB pathway in LPS-induced RAW264.7 cells. The expression of pro-inflammatory mediators and adhesion molecules are also decreased by DHMPC. Actin elongation is attenuated by inactive form of cofilin which is phosphorylated by LIMK 1/2. Our studies demonstrated that the protective effects of DHMPC in inflammation of LPS-induced RAW264.7 and SVEC4-10 cells.