2026-11-092026-11-09https://scholars.csmu.edu.tw/handle/123456789/38789Diabetic nephropathy is a major mortal cause in patients with end stage diabetes. This disorder is associated with high glucose increasing the production of reactive oxygen species (ROS) or ROS-derived oxidized low density lipoprotein (oxLDL). Previous studies showed that ROS play a role in pathogenesis and pathophysiological mechanisms triggering diabetic nephropathy. In our preliminary data, in diabetic rats, we also find the phenomena as following: (1) showing hydropic change in kidney specimen (HE stain), (2) showing thickness in glomerular basement membrane (Electron microscope examination), (3) high LDL and high triglyceride in plasma, (4) increasing lipid peroxidation in kidney specimen, (5) reducing GSH and catalase activity in kidney specimen. According to the above, in diabetes, the usage of antioxidative materials might elevate antioxidation status and reduce oxLDL to decelerate the development of diabetic nephropathy. Base on the previous studies, apple polyphenol possesses anti-cancer, anti-oxidation, anti-inflammation, and oxLDL-induced oxidative damage and fatty acid oxidation. In this study, we will use apple polyphenol to address this issue. We assume that apple polyphenol possesses the ability to reduce high glucose induced ROS and oxLDL, to reduce oxidative damage and mesangial cell damage, and further to ameliorate diabetic nephropathy. The followings will be included in our study. 【First year】 We will use streptozotocin to induce type 1 diabetes in SD rat to clarify the effect of apple polyphenol in diabetic nephropathy. The blood sample will be collected to detect the blood biochemical assay including sugar, BUN, creatinine, triglyceride, LDL, HDL, GOT, GPT, and oxidative markers including GSH-Px, SOD, catalase, MDA. The kidney specimen will be collected to examine the morphological change, thickness of glomenular basement membrane, oxidative markers, and protein expressions of TGF-β, NF-κB, PKC, Akt, Bcl-2, Bax. 【Second year】 We will use high glucose to treat mesangial cell to clarify the mechanism of apple polyphenol. As pre-treating with apple polyphenol, the cell will be treated with 25 mM of glucose to examine the ROS generation, catalase activity, SOD activity, cell death and the related proteins, such as bax, bcl-2, bad, p53, PKC, casp3, cyt c, and the change of extracellular matrix molecules. 【Third year】 We will use oxLDL to treat mesangial cell to clarify the mechanism of apple polyphenol. As pre-treating with apple polyphenol, the cell will be treated with 100 μg/ml of oxLDL to examine the ROS generation, TBARs, catalase activity, SOD activity, cell death and the related proteins, such as bax, bcl-2, bad, p53, LOX-1, and the change of extracellular matrix molecules. Taken the above together, we predict apple polyphenol would possess the effects including: (1) decelerating diabetic nephropathy, (2) decreasing high glucose-induced mesangial cell damage via reducing cell death or changing the composition of extracellular matrix, (3) decreasing oxLDL-induced mesangial cell damage via reducing cell death or changing the composition of extracellular matrix. According to those, the apple polyphenol would be developed to a health-caring food in ameliorating diabetic nephropathy.糖尿病腎病變為末期糖尿病患之主要死因，在高血糖環境中所誘發的氧化壓力及 衍生出之oxLDL 可能是促進併發症的主要機轉。除了先前文獻的研究資料顯示糖尿 病腎的致病機轉外，在我們預做的實驗中，也發現糖尿病引起腎臟病變的現象如下： (1) 在HE stain 中顯示出hydropic change；(2) 電鏡檢查中則出現腎絲球基底增厚的 現象；(3) 在血脂肪亦出現高LDL 及高TG；(4) 腎臟組織中lipid peroxidation 增加； (5) 腎臟組織中GSH 減少及catalase 活性下降。因此具有抗氧化能力的物質或許可 以藉由提升糖尿病患體內之抗氧化能力及降低oxLDL，來達到改善併發症的功效。 本研究採用之蘋果多酚為天然抗氧化物，目前已知其具有抗癌症、抗氧化及抗發炎 的能力，且對於oxLDL 之氧化傷害及脂肪酸的氧化具有抑制作用，因此我們推測蘋 果多酚應能有效清除細胞代謝高血糖所產生的過量自由基及降低oxLDL，來減少氧 化壓力與腎間質細胞的損傷，進而抑制糖尿病腎病變的形成。 以下為本研究將進行的內容: 第一年：以STZ 誘發第一型糖尿病之動物模式，來探討蘋果多酚之抗氧化能力對改 善糖尿病腎病變之影響及其機制探討。我們將收集血液測定血液生化值(包 括sugar, BUN, creatinine, triglyceride, LDL, HDL, GOT, GPT)及氧化指標 (GSH-Px, SOD, catalase, MDA)；以及腎臟檢體進行電子顯微鏡檢察、HE stain、氧化指標及蛋白質檢查 (TGF-β, NF-κB, PKC, Akt, Bcl-2, Bax) 第二年：利用腎絲球細胞於高糖環境下，探討蘋果多酚的抗氧化效果及清除自由基 能力，以及高糖產生ROS 的機制探討。研究內容為預處理蘋果多酚後，以 高濃度的葡萄糖 (25 mM) 處理腎間質細胞，測定ROS generation、catalase activity 及SOD activity；high glucose 誘導細胞死亡之狀況及蛋白分析 (bax, bcl-2, bad, p53, PKC, casp3, cyt c)；high glucose 誘導細胞外基質相關分子的 改變情形。 第三年：利用腎絲球細胞於oxLDL 存在的狀況下，探討蘋果多酚抑制oxLDL 所造 成的腎絲球損傷及其相關的機轉。研究內容為預處理蘋果多酚後，以oxLDL (100 μg/ml) 處理腎間質細胞，測定ROS generation、TBARs、catalase activity 及SOD activity；high glucose 誘導細胞死亡之狀況及蛋白分析 (bax, bcl-2, bad, p53, LOX-1)；high glucose 誘導細胞外基質相關分子的改變情形。 從以上的研究結果，我們預期蘋果多酚有（1）改善糖尿病腎病變；（2）抑制high glucose-induced mesangial cell damage 是經由抑制細胞死亡或改變細胞外基質組成而 產生；（3） 抑制oxLDL-induced mesangial cell damage 是經由抑制細胞死亡或改變 細胞外基質組成而產生。因此，以上的研究可將蘋果多酚發展為一種預防糖尿病腎 病變之保健食品。蘋果多酚糖尿病腎病變氧化壓力氧化型低密度脂蛋白凋謝死亡apple polyphenoldiabetic nephropathyoxidative stressoxidizedlow density lipoproteinapoptosisProject