PEDF抑制3T3L1脂肪細胞生成及保護老鼠免於經高脂飲食誘發的肥胖及代謝性異常

Abstract

肥胖是現代人的文明病，帶來許多代謝疾病及心血管疾病，表現出高血糖、高血脂和胰島素耐受性差等代謝異常；而脂肪組織的氧化壓力和發炎是代謝疾病的成因，脂肪細胞分化和脂肪分解的動態平衡調節更是近年來研究的核心。Pigment epithelium-derived factor (PEDF)是由視網膜色素上皮細胞培養液分離出，近年來被廣為研究的蛋白質，含有促進神經生長、抑制血管生成、抑制腫瘤生長、平滑肌細胞增生及發炎等功能。本實驗目的主要探討PEDF對於抑制肥胖及抑制脂肪生成的功能及其調控機制。在動物實驗上，以C57B/6公鼠餵食高脂食物(high fat diet, HF diet)為誘發肥胖(Diet induced obesity, DIO)的動物模式，藉由HF diet合併PEDF服用及以高脂食物誘發肥胖(DIO)的小鼠再服用PEDF，來觀察PEDF對肥胖的抑制及治療效果。在細胞實驗部分，細胞實驗則是以3T3-L1脂肪前細胞分化為脂肪細胞的過程中，在不同時間加入PEDF共同作用，藉由觀察脂肪油滴生成分化的情形，評估PEDF對脂肪細胞分化抑制及脂肪油滴代謝的影響。在動物實驗結果部分發現，PEDF能有效減緩小鼠因HF誘發增加的體重、抑制及減緩脂肪的堆積，同時具有降血糖、血脂、改善胰島素耐受性及降低脂肪組織內巨噬細胞浸潤的發炎反應；另外對於褐色脂肪(Brown adipose tissue; BAT)方面，PEDF能夠增加BAT的表現，藉此提高身體代謝的功能；在肝臟方面，PEDF可以減少肝臟內脂肪堆積、纖維化及氧化性壓力(Reactive oxygen stress; ROS)。在細胞實驗結果部分，PEDF能有效減少脂肪油滴分化及生成。藉由流式細胞及西方墨點法的分析，進一步發現，PEDF可以藉由調控前脂肪細胞的細胞週期，延緩細胞由G0G1進入S時期，經此來達到抑制脂肪細胞分化及生成，同時PEDF也具有促使脂肪分解(Lipolysis)，降低脂肪細胞總量。將脂肪組織及細胞蛋白質萃取物，藉由西方墨點法的分析，結果顯示PEDF可能是透過抑制mTOR-S6K的活化，進一步抑制PPAR-γ、CEBP-α、CEBP-β等脂肪生成相關轉錄因子的表現。綜合上述的研究結果顯示，在肥胖代謝相關疾病中，PEDF可以提供未來一個新的治療選擇。

Obesity is the modern disaster of human being, and is the major cause of many metabolic diseases and cardiovascular diseases. It presents with metabolic disorders such as hyperglycemia, hyperlipidemia, and insulin resistance. Previous studies reveal that the oxidative stress and inflammation in the adipose tissues take the responsibility of metabolic diseases, and they also concentrate in the differentiation of adipocytes and the dynamic balance of lipolysis. Pigment epithelium-derived factor (PEDF), a well-known cytokine which isolate from retina pigment epithelial cells, has the multiple function of potential neuronal differentiating activity, anti-neoplasm, smooth muscle growth inhibition, and anti-inflammation. Further studies also demonstrate that the adipose tissue is not only the source of PEDF, but also the target of this unique protein. The aim of this study is to investigate the role of PEDF to inhibit obesity and anti-adipogenesis. In vivo, we feed C57BL/6 mice with high fat (HF) diet as animal model. The animals are divided into HF fed with PEDF group and DIO (diet-induced obesity) with PEDF group, compared with control group, to study the anti-obesity effect of PEDF. In vitro, we use 3T3-L1 preadipocyte as cell model, and treat with PEDF during differentiation and after differentiation. By lipid droplet formation, we inspect the inhibition effect of PEDF to adipocyte differentiation and lipid metabolism such as lipolysis and lipogenesis. In animal study, the data shows that PEDF effectively decreases body weight gain, suppresses and slow down the obesity, decreases white adipose tissue (WAT) formation and inflammation. It also improves insulin resistance, hyperglycemia and decrease serum lipid. In brown adipose tissue (BAT), PEDF decreases the lipid accumulation. In liver, our study proved that PEDF decreases lipid formation and fibrosis. In vitro study, PEDF diminishes lipid droplet formation after 3T3-L1 differentiations. We also found out that PEDF prolongs the cell cycle progress and promotes lipolysis, which may work through mTOR-S6K pathway and subsequent transcription factors such as PPAR-γ, CEBP-α, and CEBP-β. These findings prove the hypothesis that PEDF diminishes white adipocyte formation, promotes lipid metabolism, and subsequent weight loss in mice. In conclusion, PEDF protects against high-fat diet-induced obesity and metabolic disorders in mice, and inhibits adipocyte differentiation in 3T3-L1 cells. It provides a new treatment trend for obesity in the future.