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標題: | 探討Fenofibrate對於治療糖尿病視網膜病變可能之機制 Implication of Fenofibrate for Diabetic Retinopathy Treatment |
作者: | Ying-Jung Hsu 許穎絨 |
指導教授: | 楊長豪(Chang-Hao Yang) |
共同指導教授: | 楊偉勛(Wei-Shiung Yang) |
關鍵字: | 糖尿病視網膜病變,降血脂藥物fenofibrate,氧化壓力,細胞凋亡,第一型糖尿病大鼠模式,脂聯素, diabetic retinopathy,fenofibrate,oxidative stress,apoptosis,rat model of type 1 diabetes,adiponectin, |
出版年 : | 2021 |
學位: | 博士 |
摘要: | 糖尿病視網膜病變是一種重要的糖尿病微血管併發症,也是在已開發國家中造成失明的主因之一。有大型臨床研究文獻指出降血脂藥物fenofibrate 為一種 peroxisome proliferator-activated receptor-α促效劑並且具有阻止糖尿病視網膜病變病程的潛能。我們的研究包含兩大部分,第一部分是fenofibrate對於氧化壓力所引起視網膜/脈絡膜血管內皮細胞凋亡之保護作用。我們評估RF/6A 細胞在受到paraquat所引發氧化壓力情況下fenofibrate 的保護作用並研究潛在的機制。相較於paraquat刺激的細胞,預先經過fenofibrate 處理可抑制自由基的產生,降低細胞凋亡比例並且減少粒線體膜電位的改變。Fenofibrate 會使peroxiredoxin (Prx)、thioredoxins (Trxs)、B-cell lymphoma 2 (Bcl-2)及Bcl-xl 的 mRNA 增加,並降低B-cell lymphoma 2-associated X protein (Bax) 的mRNA。西方點墨法結果顯示fenofibrate 減少細胞凋亡反應是透過細胞質及粒線體的apoptosis signal-regulated kinase-1 (Ask)-Trx相關訊息傳導路徑,包括c-Jun amino-terminal kinase (JNK) 磷酸化,cytochrome c 由粒線體釋出,caspase 3活化,以及 poly (ADP-ribose) polymerase (PARP) 裂解。這些保護作用也許可歸因於fenofibrate 在RF/6A 細胞的抗氧化能力。此研究顯示fenofibrate 也許能在氧化壓力相關的眼科疾病中提供有效的附屬治療。第二部份是fenofibrate對於第一型糖尿病大鼠視網膜內脂聯素表現之影響。脂聯素是一種特殊脂肪細胞蛋白,並與增加糖尿病微血管併發症的風險有相關性。然而,目前仍不清楚脂聯素在糖尿病視網膜病變中所扮演的角色。在此研究中,我們研究脂聯素及其受體在糖尿病大鼠中的表現,以及評估在動物體及細胞中 fenofibrate 對脂聯素及其受體表現之影響。在糖尿病大鼠視網膜中,脂聯素及其受體的mRNA 和蛋白質表現量會增加,但經過fenofibrate 治療的大鼠其視網膜中脂聯素及其受體的mRNA 和蛋白質表現量會受到抑制。免疫螢光染色結果顯示脂聯素及受體1表現於血管內、 視網膜神經節以及內核層。脂聯素受體1 也強烈表現於血管內皮細胞,反之受體 2 只有少量表現於血管內皮細胞。在細胞實驗中,RGC-5細胞及RAW264.7細胞受高糖濃度刺激下脂聯素的表現量會上升,但預先經過fenofibrate 處理之細胞其脂聯素表現會受到抑制。脂聯素受體1及受體2 在RGC-5 細胞受高糖濃度刺激下表現量會上升,但預先經過fenofibrate 處理之細胞其受體表現會受到抑制。以上結果顯示脂聯素在糖尿病視網膜中可能是一種發炎反應前的中介物並在病程中扮演重要角色。Fenofibrate 在糖尿病視網膜中可能透過調節脂聯素及其受體的表現進而有效減緩糖尿病視網膜病變之病程。 Diabetic retinopathy (DR) is an important microvascular complication of diabetes and one of the leading causes of blindness in developed countries. Two large clinical studies showed that fenofibrate is a peroxisome proliferator-activated receptor-αagonist that has been shown to be capable of preventing DR progression. Our research comprehends two parts. The first part demonstrated that the effect of fenofibrate on oxidative stress-induced apoptosis in retinal/choroidal vascular endothelial cells. We evaluated the protective effects of fenofibrate on retinal/choroidal vascular endothelial cells under oxidative stress and investigated the underlying mechanisms using RF/6A cells as the model system and paraquat (PQ) to induce oxidative stress. Pretreatment with fenofibrate suppressed reactive oxygen species (ROS) production, decreased cellular apoptosis, diminished the changes in the mitochondrial membrane potential, increased the mRNA levels of peroxiredoxin (Prx), thioredoxins (Trxs), B-cell lymphoma 2 (Bcl-2), and Bcl-xl, and reduced the level of B-cell lymphoma 2-associated X protein (Bax) in PQ-stimulated RF/6A cells. Western blot analysis revealed that fenofibrate repressed apoptosis through cytosolic and mitochondrial apoptosis signal-regulated kinase-1 (Ask)-Trx-related signaling pathways, including c-Jun amino-terminal kinase (JNK) phosphorylation, cytochrome c release, caspase 3 activation, and poly (ADP-ribose) polymerase (PARP) cleavage. These protective effects of fenofibrate on RF/6A cells may be attributable to its anti-oxidative ability. Our research suggests that fenofibrate could serve as an effective adjunct therapy for ocular oxidative stress-related disorders, such as DR. The second part demonstrated that the effect of fenofibrate on adiponectin expression in retinas of streptozotocin-induced diabetic rats. Adiponectin is an adipocyte-specific protein that has been associated with increased risks of microvascular complications in diabetes; however, its role in the development of DR is largely unknown. In this study, we investigated the expression of adiponectin and its receptors in DR and evaluated the effects of fenofibrate on their expression in vivo and in vitro. The mRNA and protein levels of adiponectin and its receptors were elevated in retinas of streptozotocin-induced diabetic rats and were suppressed following fenofibrate treatment. Immunofluorescence staining demonstrated that adiponectin and adipoR1 were expressed in cells located within blood vessels, the retinal ganglion, and the inner nuclear layer. AdipoR1 was also strongly expressed in vascular endothelial cells, whereas adipoR2 was only expressed at low levels in vascular endothelial cells. The in vitro experiments showed that adiponectin expression was induced by high glucose concentrations in RGC-5 and RAW264.7 cells and was suppressed following fenofibrate treatment. AdipoR1 and adipoR2 levels in RGC-5 cells were elevated following exposure to high glucose concentrations and suppressed after fenofibrate treatment. Our results demonstrated that adiponectin may be a pro-inflammatory mediator in diabetic retinas and play an important role in the development of DR. Fenofibrate appears to modulate the expression of adiponectin and its receptors in diabetic retinas, effectively reducing DR progression. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/70993 |
DOI: | 10.6342/NTU202100719 |
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顯示於系所單位: | 臨床醫學研究所 |
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