陽離子固態脂質奈米粒子對輸送沙奎那維穿透血腦屏障的效應：非共價鍵調節之自組裝脂質層

Cationic solid lipid nanoparticles containing assembled lipid layer with noncovalent mediation for delivering saquinavir across the blood–brain barrier

王正欽(Cheng-Chin Wang)

指導教授 ： 郭勇志

國立中正大學/工學院/化學工程學系/博士(2013年)

摘要

本研究以可可脂(cacao butter)與膽固醇(cholesterol)作為脂質主體，stearylamine (SA)與esterquat 1 (EQ 1)為陽離子脂質，製備新穎表面的陽離子固態脂質奈米粒子(cationic solid lipid nanoparticles, CSLNs)，分析表面分子間的非共價性交互作用(noncovalent interactions)、脂質主體和自組裝脂質層之離子基團對CSLNs電泳遷移率、界達電位及粒徑的影響，並結合Ohshima軟球理論，估算CSLNs表面電荷密度。另一方面，以CSLNs作為沙奎那維(saquinavir, SQV)的輸送載體，探討脂質組成對SQV包覆率的影響，分析在不同pH值與脂質組成下SQV的釋放動力學變化，並與人腦微血管內皮細胞(human brain-microvascular endothelial cells, HBMECs)共培養，測定細胞毒性。此外，以HBMECs和星形細胞(astrocytes)共培養建立體外血腦屏障(blood–brain barrier, BBB)模型，分析CSLNs包覆SQV (SQV-CSLNs)穿透BBB模型的穿透率。研究結果顯示，CSLNs的平均粒徑介於88 ± 9 nm到275 ± 16 nm之間，且粒徑會隨著膽固醇與EQ 1重量百分比的增加而變小。經由X光光電子能譜儀(X-ray photoelectron spectroscopy, XPS)與傅立葉轉換光譜儀(Fourier-transform infrared spectroscopy)的研究證實，膽固醇與EQ 1具有強烈的非共價性交互作用。此外增加膽固醇與EQ 1的重量百分比，有助於提升CSLNs的電泳遷移、界達電位與固定電荷密度。EQ 1的重量百分比增加，會降低SQV的包覆率，當脂質主體添加25% (w/w)的膽固醇，會提升SQV的包覆率，但當膽固醇提高到75% (w/w)時，會得到相反的結果。膽固醇有助於延長SQV的釋放。HBMECs 存活率隨膽固醇重量百分比增加而降低，當EQ 1的重量百分比增加，有助於提升HBMECs的存活率。此外，SQV-CSLNs對BBB的穿透率會隨著膽固醇和EQ 1重量百分比的增加而增加。由螢光染色證實，SQV-CSLNs能被HBMECs攝入。

關鍵字

沙奎那維 ； 藥物釋放 ； 電泳遷移率 ； 陽離子固態脂質奈米粒子 ； 人腦微血管內皮細胞 ； 星形細胞

並列摘要

Innovated cationic solid lipid nanoparticles (CSLNs) containing cacao butter, cholesterol, stearylamine, and esterquat 1 (EQ 1) were fabricated. The effect of noncovalent interactions, lipid matrix and surface molecular composition on the electrophoretic mobility, zeta potential and particle size of CSLNs were investigated. The fixed charge density on CSLNs was evaluated using Ohshima’s soft particle theory. In addition, the entrapment and release of SQV in CSLNs with the mediation of cholesterol were also studied. The cytotoxicity of CSLNs to human brain-microvascular endothelial cells (HBMECs) was assessed. The permeability across the blood–brain barrier (BBB) using SQV-loaded CSLNs (SQV-CSLNs) was estimated by an in vitro model comprising a monolayer of human brain-microvascular endothelial cells (HBMECs) with regulation of human astrocytes (HAs). The results revealed that the average diameter of CSLNs is in the range from 88 ± 9 nm to 275 ± 16 nm, and decreased when the weight percentage of cholesterol and EQ 1 increased. X-ray photoelectron spectroscopy and Fourier-transform infrared spectroscopy evidenced strong noncovalent interactions between cholesterol and EQ 1. In addition, an increase in the weight percentage of cholesterol and EQ 1 led to an increase in electrophoretic mobility, zeta potential, and fixed charge density of CSLNs. The results also indicated that an increase in the weight percentage of EQ 1 reduced the entrapment efficiency of SQV. The entrapment efficiency of SQV enhanced, when the weight percentage of cholesterol increased from 0% to 25% (w/w). The reverse was true when cholesterol increased from 0% to 75% (w/w). When the weight percentage of EQ 1 increased, the viability of HBMECs enhanced. An increase in the weight percentage of cholesterol reduced the viability of HBMECs. An increase in the weight percentage of cholesterol and EQ 1 also enhanced the BBB permeability of SQV and promoted the uptake of SQV-CSLNs by HBMECs. Moreover, the fluorescent staining demonstrated that SQV-CSLNs could be internalized by HBMECs.

並列關鍵字

saquinavir ； drug release ； electrophoretic mobility ； cationic solid lipid nanoparticles ； human brain-microvascular endothelial cells ； astrocytes

參考文獻

17. Y. C. Kuo, C. Y. Chung, “Transport of zidovudine- and lamivudine-loaded polybutylcvanoacrvlate and methylmethacrylate-sulfopropylmethacrylate nanoparticles across the in vitro blood-brain barrier: characteristics of the drug-delivery system,” J. Chin. Inst. Chem. Engrs., 36, 627–638, 2005.

1. C. D. Mathers, D. Loncar, “Updated projections of global mortality and burden of disease, 2002–2030 data sources, methods and results,” World Health Organization, Geneva, 2005.

2. D. A. Balazs, W. Godbey, “Liposomes for use in gene delivery,” J. Drug Deliv., 2011, 326497, 2011.

3. S. B. Tiwari, M. M. Amiji, “A review of nanocarrier-based CNS delivery systems,” Curr. Drug Deliv., 3, 219–232, 2006.

4. D. Pissuwan, T. Niidome, M. B. Cortie, “The forthcoming applications of gold nanoparticles in drug and gene delivery systems,” J. Control. Release, 147, 65–71, 2011.