研究秦皮素對於腎臟纖維化之角色

腎臟纖維化是腎臟長期損傷且不可逆的過程，因腎功能下降導致慢性腎臟病(Chronic Kidney Disease, CKD)。當腎臟組織反覆發生發炎反應、發炎細胞浸潤以及釋放大量的傷害性分子，例如：ROS、纖維化以及細胞發炎因子，導致細胞外基質過度堆積最終造成腎臟結構或功能異常。因此釐清腎臟纖維化過程以及遏制纖維化的進程，將會是現今待解決的重要議題。秦皮素(Fraxetin)在過去研究指出具有抗氧化以及清除自由基的功能，更有研究指出Fraxetin具有抗肝臟纖維化的功效，因此本研究主要以動物以及細胞模式探討Fraxetin對於抗腎臟纖維化的作用機轉。首先我們使用單側輸尿管結紮手術(UUO)模擬小鼠發生腎臟纖維化過程，並於手術後每天餵食Fraxetin (40 mg / kg /天)處理7天以及14天。接著將老鼠犧牲並取出腎臟，再以蘇木精-伊紅染劑 (H.E. stain)染色發現，UUO組別的腎臟組織間質空洞明顯較於控制組別增加。後續以Western blotting和免疫組化染色發現UUO組別的纖維化指標蛋白α-SMA、COL4A6 (Collage type IV)以及vimentin蛋白表達增高。然而與單獨使用UUO組別相比，Fraxetin顯著減緩UUO誘導α-SMA，COL4A6和vimentin 的表達。此外，Fraxetin也證實降低UUO刺激的MAPK/ERK (Extracellular signal-regulated kinases)磷酸化。同時細胞實驗部分，我們使用老鼠腎絲球細胞(SV40 MES 13)處理IS (Indoxyl sulfate)內毒素模擬腎臟慢性損傷模式，結果發現SV40細胞處理IS內毒素後會誘導細胞加速爬行，然而fraxetin則有效減緩細胞爬行的速度。綜合動物以及細胞模式結果發現，Fraxetin 顯著減輕UUO或IS誘導的腎臟纖維化過程，因此Fraxetin 可能作為慢性腎臟疾病的潛在抗纖維化的治療方式。

Long-term kidney damage will cause the kidney tissue to repeatedly activate the inflammation response. The infiltrated inflammatory cells will release a large number of noxious molecules, such as ROS, fibrosis inducing-factor and pro-inflammation cytokine. This, in turn, will induce the accumulation of extracellular matrix. When renal fibrosis reaches the point of no return, normal renal function will decline and eventually causes chronic kidney disease (CKD). Therefore, inhibiting the process of fibrosis is an urgent problem to be solved in current medical nephrology. Studies have shown that Fraxetin not only possesses anti-oxidation and free radical scavenging property but also anti-liver fibrosis ability. In this regard, we aim to explore whether Fraxetin possesses anti-renal fibrosis effect and the underlying mechanism through animal and cell models. First, unilateral ureteral obstruction (UUO) surgery was performed to simulate the kidney fibrosis process in mice. Fraxetin was then given orally (40 mg/kg/day) for 7 and 14 days post-surgery. Mice were then sacrificed to harvest the kidneys for further analysis. Hematoxylin and eosin staining showed that UUO significantly increased the interstitial voids in the kidney tissue. Subsequent immunohistochemistry staining and western blotting revealed that UUO increased the expression of fibrosis markers such as α-SMA and COL4A6 (Collagen type IV) and the expression of vimentin. Furthermore, Fraxetin significantly inhibited the expression of α-SMA, COL4A6 and vimentin induced by UUO. In addition, Fraxetin also reduced the UUO-induced ERK (Extracellular signal-regulated kinases) phosphorylation. On the other hand, IS (Indoxyl sulfate) treatment was used to simulate the chronic kidney damage in mouse glomerulus cells (SV40). Our results showed that IS treatment will enhance the migration ability of SV 40 cells and fraxetin significantly inhibited IS-enhanced mobility. Taken together, our data showed that Fraxetin significantly reduced UUO or IS-induced renal fibrosis, indicating the potential of Fraxetin as an anti-fibrotic agent for chronic kidney disease.