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本文报导了制备包含RNA的单层脂质体(Unilamellar Liposome又名脂质小泡Vesicle)和复层脂质体(Muhilamellar Liposome,或简称脂质体Liposome)的两种新方法。一是在高压下(200公斤/厘米~2)将脂类通过0.9毫米的微孔喷入RNA水溶液;另一是通过浮力将脂类均匀地慢速(每分钟15～25微升)注入RNA溶液。经过超离心和DEAE-Sephadex A_(50)柱层析可分离和纯化单层和复层脂质体。经电镜鉴定,所得样品是典型的脂质体。单层脂质体直径在500 A以下。复层脂质体呈不均一颗粒,最大者为1微米。电镜和同位素参入均证明上述两种脂质体包裹了RNA。我们还制备了博莱霉素A_5琥珀酰硬脂酰胺的钴螫合物(Co-BLS)以参入脂类,并试图作为脂质体的“导向”装置。我们又制备了各种类型包裹~(125)Ⅰ-RNA的脂质体。经核糖核酸酶(RNase)温浴,证明脂质体可保护所包裹的~(125)Ⅰ-RNA免受RNase的降解。同时证明脂质体腹腔内注射后可促进所包的~(125)Ⅰ-RNA参入小鼠腹水型肝癌细胞,而且含Co-BLS的脂质体促进作用更为明显。通过带瘤小鼠静脉内注射,脂质体所包含的RNA可参入皮下型移植性肝癌细胞,但参入比例不如腹腔注射明显,然而肝脾内比放射性百分比并不高,与文献报导不同。上述实验结果提示了以脂质体包裹核酸或其他药物,应用于遗传工程、遗传操纵或肿瘤化疗的可能性。
This article reports two new methods for preparing RNA-containing monolayer liposomes (Unilamellar Liposome, also known as Liposomal Vesicle) and multilamellar liposomes (Muhilamellar Liposome, or Liposomal Liposome). One is to inject lipids into the aqueous RNA solution through a 0.9 mm microwell under high pressure (200 kg/cm2); the other is to buoyantly inject RNA into the RNA at a uniform rate of 15 to 25 microliters per minute. Solution. Monolayer and multilamellar liposomes can be separated and purified by ultracentrifugation and DEAE-Sephadex A_(50) column chromatography. The resulting sample was identified by electron microscopy as a typical liposome. Monolayer liposomes have a diameter of 500 A or less. Multilayer liposomes are heterogeneous particles, the largest one is 1 micron. Electron microscopy and isotope incorporation demonstrated that the two liposomes encapsulated RNA. We also prepared a cobalt conjugate of bleomycin A-5 succinylstearamide (Co-BLS) for inclusion in lipids and attempted to act as a “guide” device for liposomes. We also prepared various types of liposomes encapsulating ~125 I-RNA. The RNase bath proved that the liposomes protected the 125I-RNA from degradation by RNase. At the same time, it was proved that the intraperitoneal injection of liposomes promoted the incorporation of ~(125)I-RNA into mouse ascitic hepatoma cells, and the promotion of liposomes containing Co-BLS was more obvious. Through intratumoral injection of tumor-bearing mice, RNA contained in liposomes can be incorporated into subcutaneous transplanted hepatoma cells, but the proportion of inoculation is not as significant as intraperitoneal injection. However, the percentage of specific radioactivity in liver and spleen is not high, which is different from the literature reports. The above experimental results suggest the possibility of using liposomes to encapsulate nucleic acids or other drugs for genetic engineering, genetic manipulation or tumor chemotherapy.