儲鐵蛋白透過P-Selectin Glycoprotein Ligand-1及Grp78調控 巨噬細胞遷移及第二型糖尿病大鼠胰島素耐受性

Effects of iron storage molecules on P-Selectin Glycoprotein Ligand-1 mediated macrophage migration and type 2 diabetic rat model

王齊梅(Chi-Mei Wang)

指導教授 ： 張榮素

臺北醫學大學/公共衛生暨營養學院/保健營養學研究所/碩士(2013年)

摘要

血清中儲鐵蛋白質 (serum ferritin) 反映體內鐵質儲存量。根據近年來的公共衛生研究指出，血清中儲鐵蛋白質與肥胖、代謝症候群及第二型糖尿病發生率具相關性，但其機制目前並不清楚。巨噬細胞 (macrophage) 在鐵質代謝、細胞遷移及發炎反應中扮演重要的角色。本次研究目的為探討儲鐵蛋白質對巨噬細胞活化及遷移的相關分子機轉。細胞實驗使用儲鐵蛋白 (ferritin)、脫鐵儲鐵蛋白 (apoferritin) 及鐵質 (ferric iron) 介入小鼠巨噬細胞RAW 264.7，培養於含10％胎牛血清之Dulbecco's Modified Eagle Medium培養液中，置於37℃、含5％ CO2的環境下，收集細胞及培養液進行分析。動物實驗使用streptozotocin (STZ) 誘發雄性Wistar大鼠之第二型糖尿病，誘發成功後餵食控制飲食及富含鐵質之飲食10週，收集血清及臟器進行分析。結果顯示：(1) 1000 pM之儲鐵蛋白及脫鐵儲鐵蛋白皆可誘發RAW 264.7細胞分泌TNF-α及細胞遷移指標P-selectin glycoprotein ligand-1 (PSGL-1) (all p < 0.01)；(2) 添加儲鐵蛋白及脫鐵儲鐵蛋白皆可誘發RAW 264.7細胞遷移，儲鐵蛋白添加濃度與細胞內鐵質含量呈正相關而脫鐵儲鐵蛋白則無且儲鐵蛋白誘發之細胞遷移與鐵質含量呈現負相關 (r = -0.646; p < 0.01)；(3) 添加PSGL-1阻抗劑 10及24小時後，抑制由脫鐵儲鐵蛋白所誘發之細胞遷移 (p < 0.001 and < 0.05; respectively)，但TNF-α阻抗劑則較無效；(4) 添加Grp78阻抗劑抑制了脫鐵儲鐵蛋白所誘發之細胞遷移 (p < 0.0001)；(5) 餵食第二型糖尿病大鼠富含鐵質之飲食 (0.5 g及1 g ferric iron / kg diet) 與餵食正常飲食相比會加劇胰島素阻抗性 (p < 0.01) 及增加血漿中soluble PSGL-1濃度 (p < 0.05)。總結：正常生理濃度之儲鐵蛋白及脫鐵儲鐵蛋白為促發炎因子，可誘發巨噬細胞活化及細胞遷移，而鐵質所誘發的細胞遷移需透過Grp78及PSGL-1相關路徑調控。此外，過多鐵質亦會加劇第二型糖尿病大鼠胰島素阻抗性。

關鍵字

第二型糖尿病 ； 儲鐵蛋白質 ； 巨噬細胞 ； 遷移指標P-selectin glycoprotein ligand-1 (PSGL-1)

並列摘要

Epidemiological studies indicate elevated serum ferritin (SF) concentrations are independently associated with type 2 diabetes (T2DM). However, the underlying mechanisms, which initiate the initial processes, remain largely unknown. Macrophages play a key role in iron metabolism, host response and the initial inflammatory responses. The goal of this study was to dissect the molecular targets of SF with special focus on macrophage response. The mouse macrophage-like cell line RAW 264.7 cells were treated with various iron stimuli (SF, apoferritin and ferric iron) and cellular responses were studied. Effects of iron rich diet on streptozotocin (STZ) induced type 2 diabetic rat model were evaluated at post 10 weeks intervention. Our results showed: (1) the presence of 1000 pM SF and apoferritin triggered TNF-α and soluble P-selectin glycoprotein ligand-1 (PSGL-1) secretion in RAW 264.7 cell ( all p < 0.01); (2) SF and apoferritin induced macrophage migration but migration rate was inversely associated with intracellular iron concentrations (r = -0.646; p < 0.01); (3) PSGL-1 neutralization abrogated apoferritin-mediated RAW 264.7 cells migration, particularly at 10 and 24 hours post-stimulation (p < 0.001 and < 0.05; respectively); (4) Inhibition of Grp78 protein attenuated in apoferritin-mediated macrophage migration; and (5) High iron sensitized type 2 diabetic rats to insulin resistance in rats fed with iron rich diet compared to diabetic rats received control diet (p < 0.01). In conclusion, our preliminary results suggest iron storage molecules (ferritin and apoferritin) are pro-inflammatory molecules and excess iron may promote insulin resistance in type 2 diabetic rat model.

並列關鍵字

type 2 diabetes (T2DM) ； serum ferritin (SF) ； macrophages ； P-selectin glycoprotein ligand-1 (PSGL-1)

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