Incorporation of Farnesol Significantly Increases the Efficacy of Liposomal Ciprofloxacin against Pseudomonas aeruginosa Biofilms in Vitro
Incorporation of Farnesol Significantly Increases the Efficacy of Liposomal Ciprofloxacin against Pseudomonas aeruginosa Biofilms in Vitro
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DOI：
10.1021/acs.molpharmaceut.6b00360
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发表时间：
2016-08-01
影响因子：
4.9
通讯作者：
Smyth, H. D. C.
中科院分区：
文献类型：
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作者：
Bandara, H. M. H. N.;Herpin, M. J.;Smyth, H. D. C.
由于抗生素耐药性的迅速发展，消除囊性纤维化肺等铜绿假单胞菌感染的挑战仍然没有改变。药物对致密铜绿假单胞菌生物膜的渗透不良在感染的无效清除中起着至关重要的作用。因此，需要重新审视目前针对铜绿假单胞菌生物膜的抗生素疗法，并需要发明替代的抗生物膜策略。真菌群体感应分子 (QSM)、金合欢醇似乎对铜绿假单胞菌具有有害影响。因此，本研究旨在联合递送天然存在的 QSM 法呢醇，并使用抗生素环丙沙星作为脂质体制剂来根除铜绿假单胞菌生物膜。采用脱水-再水化方法制备四种不同的脂质体（含环丙沙星和法呢醇，Lcip+far；含环丙沙星，L-cip；含法呢醇，L-far；对照，L-con）并进行表征。通过光谱测定法和高效液相色谱法（HPLC）评估药物的包封和释放。使用标准生物膜测定评估脂质体的功效。通过 XTT 还原测定和结晶紫测定对脂质体处理的 24 小时铜绿假单胞菌生物膜进行定量评估，并通过共焦激光扫描显微镜 (CLSM) 和透射电子显微镜 (TEM) 进行定性评估。与L-cip相比，当用法呢醇（Lcip+far）封装时，从脂质体中释放的环丙沙星比L-cip更高（3.06% vs 1.48%），而当用环丙沙星（Lcip+far）封装时，与L-far相比，法呢醇释放较低（1.81% vs 4.75%）。与游离环丙沙星相比，用 Lcip+far 或 L-cip 处理时生物膜代谢显着降低（XTT，P < 0.05）。当以 Lcip+far 形式给药时，与游离环丙沙星或 L-cip 相比，实现类似生物膜抑制所需的环丙沙星浓度分别低 125 倍或 10 倍 (P < 0.05)。 CLSM 和 TEM 证实，与其他脂质体制剂相比，用 Lcip+far 处理时，生物膜主要被破坏，死细胞比例更高，生物膜杀伤深度增加。因此，法尼醇和环丙沙星的共同递送可能是通过在显着较低的抗生素剂量下增强生物膜杀灭来对抗抗生素耐药性铜绿假单胞菌生物膜的有前途的方法。
The challenge of eliminating Pseudomonas aeruginosa infections, such as in cystic fibrosis lungs, remains unchanged due to the rapid development of antibiotic resistance. Poor drug penetration into dense P. aeruginosa biofilms plays a vital role in ineffective clearance of the infection. Thus, the current antibiotic therapy against P. aeruginosa biofilms need to be revisited and alternative antibiofilm strategies need to be invented. Fungal quorum sensing molecule (QSM), farnesol, appears to have detrimental effects on P. aeruginosa. Thus, this study aimed to codeliver naturally occurring QSM farnesol, with the antibiotic ciprofloxacin as a liposomal formulation to eradicate P. aeruginosa biofilms. Four different liposomes (with ciprofloxacin and farnesol, Lcip+far; with ciprofloxacin, L-cip; with farnesol, L-far; control, L-con) were prepared using dehydration-rehydration method and characterized. Drug entrapment and release were evaluated by spectrometry and high performance liquid chromatography (HPLC). The efficacy of liposomes was assessed using standard biofilm assay. Liposome-treated 24 h P. aeruginosa biofilms were quantitatively assessed by XTT reduction assay and crystal violet assay, and qualitatively by confocal laser scanning microscopy (CLSM) and transmission electron microscopy (TEM). Ciprofloxacin release from liposomes was higher when encapsulated with farnesol (Lcip+far) compared to L-cip (3.06% vs 1.48%), whereas farnesol release was lower when encapsulated with ciprofloxacin (Lcip+far) compared to L-far (1.81% vs 4.75%). The biofilm metabolism was significantly lower when treated with Lcip+far or L-cip compared to free ciprofloxacin (XTT, P < 0.05). When administered as Lcip+far, the ciprofloxacin concentration required to achieve similar biofilm inhibition was 125-fold or 10-fold lower compared to free ciprofloxacin or L-cip, respectively (P < 0.05). CLSM and TEM confirmed predominant biofilm disruption, greater dead cell ratio, and increased depth of biofilm killing when treated with Lcip+far compared to other liposomal preparations. Thus, codelivery of farnesol and ciprofloxacin is likely to be a promising approach to battle antibiotic resistant P. aeruginosa biofilms by enhancing biofilm killing at significantly lower antibiotic doses.