Karyekar Chetan S, Fasano Alessio, Raje Sangeeta, Lu Ruliang, Dowling Thomas C, Eddington Natalie D
Pharmacokinetics Biopharmaceutics Laboratory, Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, 100 Penn Street, Baltimore, Maryland 21201, USA.
J Pharm Sci. 2003 Feb;92(2):414-23. doi: 10.1002/jps.10310.
The purpose of this study was to examine the ability of Zonula occludens toxin (Zot) to reversibly open tight junctions in bovine brain microvessel endothelial cells (BBMECs) to enhance drug delivery via the paracellular pathway. Transport across BBMEC monolayers was examined for molecular weight markers and chemotherapeutic agents ([(14)C]sucrose, [(14)C]inulin, [(3)H]propranolol, [(3)H]doxorubicin, and [(14)C]paclitaxel) with Zot (0.0-4.0 microg/mL). TEER of monolayers was measured to assess effect and reversibility of Zot. Cell viability of BBMEC in the presence of Zot was assessed by trypan blue exclusion staining. Apparent permeability (P(app)), enhancement ratio (R), and percent increase in transport determined were statistically compared by ANOVA. A significant increase (p < 0.05) in P(app) was observed for the transport of [(14)C]sucrose, [(14)C]inulin, [(3)H]doxorubicin, and [(14)C]paclitaxel at a 4.0 microg/mL concentration of Zot. A significant concentration-dependent decrease in TEER was observed on treatment with Zot with rapid reversal to baseline after removal. Zot (4 micro/ml) was found to be nontoxic to the BBMECs after 2 hours incubation. In conclusion, Zot increased paracellular transport across the BBMEC in a reversible, concentration-dependent manner. Modulation of paracellular transport with Zot may be used to increase the brain permeability of potent central nervous system-active drugs, including anticancer agents.
本研究的目的是检测闭合蛋白毒素(Zot)可逆性打开牛脑微血管内皮细胞(BBMECs)紧密连接以增强经细胞旁途径药物递送的能力。使用Zot(0.0 - 4.0μg/mL)检测了分子量标记物和化疗药物([¹⁴C]蔗糖、[¹⁴C]菊粉、[³H]普萘洛尔、[³H]阿霉素和[¹⁴C]紫杉醇)跨BBMEC单层的转运。测量单层的跨上皮电阻(TEER)以评估Zot的作用和可逆性。通过台盼蓝排斥染色评估Zot存在下BBMEC的细胞活力。通过方差分析对测定的表观渗透率(P(app))、增强率(R)和转运增加百分比进行统计学比较。在Zot浓度为4.0μg/mL时,观察到[¹⁴C]蔗糖、[¹⁴C]菊粉、[³H]阿霉素和[¹⁴C]紫杉醇转运的P(app)显著增加(p < 0.05)。在用Zot处理后观察到TEER有显著的浓度依赖性降低,去除后迅速恢复到基线。孵育2小时后发现Zot(4μg/ml)对BBMEC无毒。总之,Zot以可逆的、浓度依赖性方式增加了跨BBMEC的细胞旁转运。用Zot调节细胞旁转运可用于增加强效中枢神经系统活性药物(包括抗癌药物)的脑通透性。