Department of Medical Biochemistry and Global COE program, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526, Japan.
J Control Release. 2010 Jan 25;141(2):208-15. doi: 10.1016/j.jconrel.2009.09.019. Epub 2009 Oct 6.
Liposomal amphotericin B (AMPH-B), also known as AmBisome, exhibits a potent antifungal effect through its binding to ergosterol contained within the fungal cell membrane. However, the mechanism responsible for the movement of AmBisome-derived AMPH-B to the fungal cell membrane through the cell wall is not yet clear. Therefore, in the present study we aimed at elucidating this mechanism operating in Saccharomyces cerevisiae. AmBisome showed its antifungal effect against S. cerevisiae at 35 degrees C but not at 4 degrees C, whereas free AMPH-B was effective at both temperatures. A significant difference in the amount of AMPH-B transferred to the fungal cells between incubation at 4 and 35 degrees C was also observed when AmBisome was used. Confocal microscopic study, however, indicated that NBD-labeled AmBisome was localized on the surface of the fungal cells at either temperature. To decrease the affinity of AMPH-B for the liposomal membrane, we entrapped AMPH-B in fluid liposomes containing egg yolk phosphatidylcholine (EPC) instead of hydrogenated soy PC (HSPC). These liposomes showed the antifungal effect even at 4 degrees C. On the contrary, AMPH-B in liposomes containing ergosterol (Erg-AmB) instead of cholesterol showed a significantly weaker antifungal effect at 35 degrees C with reduced transfer of AMPH-B to the fungal cells. These results suggest that not the binding of AmBisome to target cells but the transfer of AMPH-B from liposomal membrane of AmBisome to the cell membrane is critical for the antifungal activity of AmBisome. This transfer is dependent on the temperature, fluidity of the liposomal membrane, and the affinity of AMPH-B for the fungal cell membrane.
脂质体两性霉素 B（AMPH-B），也称为两性霉素 B 脂质体，通过与真菌细胞膜内的麦角固醇结合发挥强大的抗真菌作用。然而，通过细胞壁将两性霉素 B 脂质体衍生的 AMPH-B 转运到真菌细胞膜的机制尚不清楚。因此，本研究旨在阐明两性霉素 B 脂质体在酿酒酵母中发挥作用的机制。两性霉素 B 脂质体在 35°C 时对酿酒酵母表现出抗真菌作用，但在 4°C 时无效，而游离的 AMPH-B 在这两种温度下均有效。当使用两性霉素 B 脂质体时，还观察到在 4°C 和 35°C 孵育时 AMPH-B 转运到真菌细胞的量有显著差异。然而，共焦显微镜研究表明，在两种温度下，NBD 标记的两性霉素 B 脂质体都定位于真菌细胞的表面。为了降低 AMPH-B 与脂质体膜的亲和力，我们将 AMPH-B 包封在含有卵黄卵磷脂（EPC）而不是氢化大豆卵磷脂（HSPC）的流体脂质体中。这些脂质体即使在 4°C 时也表现出抗真菌作用。相反，含有麦角固醇（Erg-AmB）而不是胆固醇的脂质体中的 AMPH-B 在 35°C 时显示出明显较弱的抗真菌作用，并且 AMPH-B 向真菌细胞的转移减少。这些结果表明，不是两性霉素 B 脂质体与靶细胞的结合，而是两性霉素 B 脂质体从脂质体膜向细胞膜的转移对于两性霉素 B 脂质体的抗真菌活性至关重要。这种转移取决于温度、脂质体膜的流动性以及 AMPH-B 与真菌细胞膜的亲和力。
