Preparation and characterization of ciliary neurotrophic factor sustained-release microcapsules

doi:10.3969/j.issn.2095-4344.2014.34.008

Abstract

Abstract:

BACKGROUND: Conventional ophthalmic delivery of ciliary neurotrophic factor (CNTF) is extremely difficult to pass the blood-retinal barrier, resulting in a very low bioavailability and the need of long-term drug delivery. To solve the problem, the CNTF can be encapsulated in a semi-permeable membrane to form a microcapsule, which may then achieve the release of bioactive substances encapsulated, or bioactive molecules secreted by living cells and small molecular metabolites through semi-permeability of the special membrane.

OBJECTIVE: To prepare a special structural CNTF sustained-release microcapsule.

METHODS: A selected poly(ether sulfone) hollow fiber was cut into 1 cm long with its two ends sealed by 1181-M medical adhesive using UV curing. To prepare CNTF encapsulated microcapsule, one end was first sealed, and then the CNTF was loaded to poly(ether sulfone) microcapsule from the other end which then was sealed. The leaching liquor of sustained-release microcapsule was co-cultured with mouse fibroblast L929, to observe the cytotoxicity of the microcapsule. The sustained-release microcapsule was co-cultured with mouse retinal pigment epithelial cells, to observe the cell adhesion ability of the microcapsule. The CNTF sustained-release microcapsule was immersed in physiological saline, to observe the degradability. Moreover, in vitro release behavior of immunoglobulin and CNTF were evaluated.

RESULTS AND CONCLSION: The CNTF sustained-release microcapsule had an inner diameter of about 398 μm and a membrane thickness of about 145 μm. The microcapsule presents a lot of macropores in the outer wall and many 10 nanometers micropores in the inner wall. The sustained-release microcapsule was not degraded in saline within 4 months, indicating good cell compatibility. The microcapsule can selectively release CNTF while protecting against invading of antibodies (IgG), showing its good selective permeability. Meanwhile, the sustained-release microcapsule improved the initial burst release of traditional drug delivery vesicles. The microcapsule presents a mild sudden release in the middle stage, and then a sustained release.

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

全文链接：

Key words: ciliary neurotrophic factor, delayed-action preparations

CLC Number:

R318

Li Xiao-li, Wang Ming-bo, Chen Chang-sheng, She Zhen-ding. Preparation and characterization of ciliary neurotrophic factor sustained-release microcapsules[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(34): 5455-5460.

Figures/Tables 2

2.1 CNTF缓释微囊微观结构形貌 CNTF缓释微囊的微观形貌如图1所示，外层由很多疏松的大孔构成(图1A，B)，内层的囊孔较密集，孔径为10 nm左右(图1C)，满足其半渗透性的孔径要求。利用ImageJ软件对10根CNTF缓释微囊100个不同位置的内径及壁厚进行测量，结果显示纤维囊的内径为(398.0±12.6) μm，壁厚(145.0±20.9) μm。本实验制备微囊的直径为Thanos等[33]报道的NT501中聚醚砜中空纤维囊直径的一半，增加了表面积/体积比。 2.2 CNTF缓释微囊的生物相容性评价阴性对照组的A值为0.634±0.091，CNTF缓释微囊的A值为0.651±0.094，由相对细胞存活率公式得细胞的存活率为102.68%，根据GB/16886标准细胞毒性评价，CNTF缓释微囊的细胞毒性为0级。利用小鼠视网膜上皮细胞黏附在CNTF缓释微囊内壁24，72 h后，对细胞进行Live-Dead染色，由图2可知活细胞在CNTF缓释微囊内壁上黏附数量明显增多，因此，CNTF缓释微囊内壁有利于促进细胞增殖、黏附生长，具有良好的生物相容性。 2.3 CNTF缓释微囊的体外降解性能为了探究CNTF缓释微囊在眼科植入体内的降解，采用生理盐水为模拟眼液研究了缓释微囊的体外降解性能。由图3降解质量曲线可知，在4个月内CNTF缓释微囊的质量前后相差不大，在误差范围内，可认为基本无降解。因此，CNTF缓释微囊具有良好的稳定性。 2.4 CNTF缓释微囊的体外释放曲线将一定量CNTF与模拟蛋白IgG分别加载至PES中空纤维膜中，考察它们的在微囊中的体外释放行为。由图4A可知，在前7 h，CNTF基本呈现零释放，这可能是由于CNTF透过内壁的微孔及数百微米的厚壁需要一定的时间所致。经历这段时间后，累积的CNTF在7-24 h内呈现短暂的突释，其后从第2天至第7天以缓和的速度平稳释放，并在第7天达到65%的释放量。而对于模拟蛋白IgG，体外释放7 d内的测试A值均与空白样的A值相接近(图4B)，可认为在测量误差范围内，IgG基本无释放，不能通过微囊内壁的微孔。因此，特制结构的聚醚砜微囊对于蛋白的释放具有选择性。"

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