奈米氧化鋅對人類臍靜脈內皮細胞發炎反應之細胞黏著分子作用機制探討

奈米科技產業已經是目前全世界感興趣且著手發展的科技領域之一。因為奈米粒子的大小皆低於100 nm，故此會這奈米物質會產生新的物理化學性質不同於以往的較大顆粒的非奈米物質。然而此種新穎的奈米性質卻也會導致其他未知的毒性，特別關於心血管的毒性，故此我們研究奈米氧化鋅影響人類臍靜脈內皮細胞(HUVECs)之發炎反應機制。在體外試驗中，利用細胞毒性試驗，結果發現在10 μg/ml的濃度下，不論是奈米氧化鋅或是非奈米氧化鋅對於細胞存活率差異並不大，但在利用黏著試驗中，可發現將HUVECs處理奈米氧化鋅24小時後，會有劑量–效應現象去增加THP-1的細胞黏著於HUVECs，其中細胞黏著分子ICAM-1的蛋白質與mRNA確實會被奈米氧化鋅誘導出來且有劑量–與時間–效應的現象。我們也發現奈米氧化鋅(10 μg/ml)會誘導JNK的磷酸化與ROS的生成，然而SP600125 (JNK抑制劑)確實可以減少奈米氧化鋅所造成的ICAM-1累積，但是ROS產生則與ICAM-1表現並無相關性。而奈米氧化鋅也會導致細胞中Rac1/cdc 42/MLK3訊息傳遞路徑的活化，而ICAM-1則可以藉由NSC23766 (Rac1/cdc 42抑制劑)減少其表現量。此外在體內試驗當中，將apoE基因剔除小鼠連續餵食奈米氧化鋅十三天後，利用免疫組織染色觀察胸主動脈，其中ICAM-1的表現與動脈粥狀硬化的發展是有關聯性的，實驗結果確實也發現在胸主動脈之ICAM-1染色有明顯的改變。故此推測機制可能是奈米氧化鋅藉由Rac1/cdc 42/MLK3促使JNK磷酸化導致ICAM-1的表現，且奈米氧化鋅會導致apoE基因剔除小鼠動脈中ICAM-1累積可能會形成動脈粥狀硬化。

關鍵字

奈米氧化鋅；發炎反應；細胞黏著分子；奈米粒子；動脈粥狀硬化

並列摘要

Nanotechnology is dynamically developing fields of scientific interest in the entire world. Because nanoparticles (NPs) are sized below 100 nm, they have the new physical-chemical characteristics of nano-sized materials and differ substantially from those of bulk materials. However, the novel properties of NPs lead to lots of adverse effects, especially in cardiovascular toxicity. Therefore, we aim to investigate that the effect of NP ZnO on inflammation response in human umbilical vein endothelial cells (HUVECs). In vitro study, the results showed that the cell viability measured by MTT assay was no different compared of 10 μg/ml NP ZnO treatment and non-NP ZnO treatment. In adhesion molecule functional test by using adhesion assay, THP-1 cells were adhered to HUVEC by NP ZnO (0.1-10 μg/ml) in dose-dependent manner in 24 hr. NP ZnO could significantly induce the expression of ICAM-1 protein and mRNA in HUVECs in a dose- and time-dependent manner. We also found that NP ZnO (10 μg/ml) would cause the phosphorylation of JNK and ROS generation. Furthermore, SP600125 (JNK inhibitor) reduced NP ZnO-induced ICAM-1 accumulation. However, the generation of ROS and ICAM-1 expression was not correlated. Then, NP ZnO treatment could activate the ICAM-1 expression through Rac1/cdc 42/MLK3 signaling pathway. And the expression of ICAM-1 could be inhibited by NSC23766 (Rac1/cdc 42 inhibitors). In vivo study, we applied immunohistochemistry to stain thoraic aorta of apoE-deficient mice after fed with NP ZnO continuously lasting to 13 days. ICAM-1 staining was detected in thoraic aorta, and the level of ICAM-1 has been shown to be correlated with the progression of atherosclerosis. In summary, (1) It suggests that the phosphorylation of JNK participates in NP ZnO-induced ICAM-1 expression in HUVEC, while Rac1/cdc 42/MLK3 plays a role in this mechanism. (2) Our results support the ZnO-induced ICAM-1 accumulation may cause the formation of atherosclerosis in the artery area in apoE-deficient mice.

並列關鍵字

NP ZnO ； nanoparticles ； inflammation response ； ICAM-1 ； JNK ； c-jun ； MLK3 ； Rac1/cdc42 ； atherosclerosis

參考文獻

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奈米氧化鋅對人類臍靜脈內皮細胞發炎反應之細胞黏著分子作用機制探討

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