课题背景：本课题获国家自然科学基金资助，项目名称：多聚微囊化转脑啡肽基因细胞的镇痛效应和免疫耐受研究，项目批准号：39900139。①内容：脑啡肽基因修饰细胞的制备与微囊化处理；微囊化细胞的存活情况、分泌功能、致瘤性及免疫耐受研究；微囊化细胞用于慢性疼痛的研究。②成果：细胞移植前的微囊化处理，使宿主接受移植后的免疫反应明显减轻；微囊小体对移植细胞有明显的免疫保护作用，移植细胞在宿主蛛网膜下腔的存活和效应时间明显延长；转染人前脑啡肽原基因的体细胞在Wistar大鼠具有和人嗜铬细胞等效的椎管腔镇痛效应，可以取代嗜铬细胞而成为操作性更强的镇痛用细胞；微囊包裹对细胞的分泌功能和致瘤性无显著性影响。

相关链接：对于慢性疼痛的治疗，药物镇痛和生物细胞镇痛是研究的热点，生物细胞在成瘾性和依赖性方面优于药物，但其存活时间和移植后的免疫排斥反应是需要解决的问题。转染人前脑啡肽原基因细胞的体细胞可在蛛网膜下腔长期存活并发挥镇痛效应，但转染效率如何、是否可取代嗜铬细胞而成为操作性更强的镇痛用细胞还在进一步研究中。

偏倚或不足：实验设计中未比较海藻酸钠-聚赖氨酸-海藻酸钠和单纯的海藻酸钠微囊这两种微囊对人嗜铬细胞、小鼠黑色素瘤细胞（B16-F1）及对照细胞小鼠成纤维细胞(NIH3T3)细胞数量、活性的影响。

摘要
目的: 观察海藻酸钠-聚赖氨酸-海藻酸钠（APA）微囊对囊内细胞的存活、生长及凋亡的影响，以验证免疫隔离APA微胶囊的生物膜性和生物活性。
方法: 实验于2004-01/05在华中科技大学同济医学院附属同济医院麻醉科实验中心完成。取人嗜铬细胞原代培养，小鼠黑色素瘤细胞和小鼠成纤维细胞传代培养，将APA微囊分别包裹上述3种细胞并进行培养；根据3种细胞微囊化与否分成6组，利用MTT比色法检测各组细胞存活和生长情况，共观察9 d，并用流式细胞仪检测各组细胞凋亡情况。
结果: ①在实验观察的9 d内，人嗜铬细胞、小鼠黑色素瘤细胞和小鼠成纤维细胞可在微囊内以细胞团的形式生长、存活。②MTT比色法检测显示各组囊内细胞与相应组裸细胞的生长、存活情况比较差异无显著性意义（P > 0.05）。③流式细胞仪检测各组囊内细胞与相应组裸细胞的凋亡百分率比较差异亦无显著性意义（P > 0.05）。
结论: 制备的APA微囊化材料及制备方法对细胞活性、生长状况及凋亡情况无不良影响，表明包裹细胞的APA微囊有良好的生物相容性和生物活性。
关键词: MTT比色法；凋亡；微囊化；嗜铬细胞；黑色素瘤细胞；生物材料

陈冀衡，杨茂元，毕好生.微囊技术对人嗜铬细胞和小鼠黑色素瘤细胞凋亡的影响[J].中国组织工程研究与临床康复，2008，12(6):1017-1021 [www.zglckf.com/zglckf/ejournal/upfiles/08-6/6k-1017(ps).pdf]

Effect of microencapsulation technique in the apoptosis of human chromaffin cells and mouse melanoma cells

Abstract

AIM：To observe the survival, growth and apoptosis of alginate-polylysine-alginate (APA) microencapsulated cells, and verify the biological membrane and activity of APA microcapsules with immunoisolation.
METHODS: The experiment was carried out in the test center, Department of Anesthesiology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology from January to May in 2004. Human chromaffin cells were harvested for primary culture, mouse melanoma cell and mouse fibroblast were subcultured, then three kinds of cells were encapsulated and cultured in APA microcapsules. According APA microencapsulation or not, three kinds of cells were divided into 6 groups and their survival and growth were measured for 9 days by MTT colorimetric assay. Flow cytometry was used to detect the cell apoptosis.
RESULTS: ①During 9-day observation, human chromaffin cells, mouse melanoma cell and mouse fibroblast in the microcapsules were all survived and grew in the clusters.②MTT colorimetric assay showed that the differences of cell growth and survival were not significant between the microencapsulated group with APA and non-microencapsulated group with APA (P > 0.05).③Flow cytometry revealed that there was no significant difference in the rate of apoptosis between the microencapsulated group and non-microencapsulated group (P > 0.05).
CONCLUSION: APA microencapsulation technique has no harm to the activity, growth and apoptosis of cells, indicating good biological compatibility and activity of APA.

Chen JH, Yang MY, Bi HS.Effect of microencapsulation technique in the apoptosis of human chromaffin cells and mouse melanoma cells.Zhongguo Zuzhi Gongcheng Yanjiu yu Linchuang Kangfu 2008;12(6):1017-1021(China)
[www.zglckf.com/zglckf/ejournal/upfiles/08-6/6k-1017(ps).pdf]