Development of a novel artificial red blood cell via membrane emulsification technique

体外构建三维组织并将其植入体内是组织工程的最终目标。为此，需要人工氧载体来为再生组织提供足够的氧气。本研究采用膜乳化法、无创化学反应法和动物细胞培养法相结合的方法，制备了一种粒径在1 ~ 5 μm之间、粒径分布窄的新型氧载体。氧载体通过减少内吞作用实现低细胞毒性，通过使用牛血红蛋白（Hb）实现高氧容量和低成本。事实上，上述各点我们都做到了。我们从牛血中提取了一种新鲜、有活性的血红蛋白，纯度达98%，经SDS-PAGE和UV-vis证实。通过加入BSA防止膜污染，我们成功地乳化了10wt%的Hb，其乳液尺寸约为100 μ m。4μm。随后用戊二醛交联得到了一种新的氧载体，其P50为19 mmHg。载体对HepG 2细胞的毒性低于同浓度Hb。

Construction of three-dimensional tissue in vitro and its implantation are an ultimate goal in tissue engineering. For this purpose, an artificial oxygen carrier is necessary to supply enough oxygen to regenerative tissues. In this research, we fabricated a new oxygen carrier which diameter is between 1 and 5 μm and has a narrow size distribution by the combination of membrane emulsification, non-invasive chemical reaction, and animal cell culture. The oxygen carrier achieved low cytotoxicity of cells by the reduction of endocytosis, high oxygen capacity, and low cost by the use of bovine hemoglobin (Hb). In fact, we achieved all the above-mentioned points. We extracted a fresh and active Hb from bovine blood by 98 % purity, which were confirmed by SDS-PAGE and UV-vis. We successfully emulsified 10 wt% of Hb by preventing the fouling of the membrane by addition of BSA, which emulsion size was ca. 4μm. Subsequent crosslinking by glutaraldehyde brought a new oxygen carrier which P50 was 19mmHg. The toxicity of the carriers to HepG2 was lower than that of the same concentration of Hb.