两段法培养适合临床移植应用的黑素细胞的安全性研究

Security of human melanocytes cultured by two step method

目的:探讨使用两段法培养黑素细胞,以消除佛波醇酯(TPA)的致瘤危险,增加培养细胞的安全性,从而适合临床移植的需要。方法:两段法培养黑素细胞:先用含TPA培养液培养纯化黑素细胞,然后用不含TPA的培养基继续扩增传代。通过细胞形态、生长特性、细胞周期等检测观察有无TPA时培养细胞的特性。接种裸鼠检测培养细胞的致瘤性。结果:用含TPA培养液容易获得纯化的黑素细胞;扩增传代时改用不含TPA的培养基,黑素细胞由细长的多分支逐渐变成两极的宽梭形。生长速度略微减慢,细胞周期显示增殖减缓。裸鼠致瘤实验阴性。结论:两段法培养黑细胞成功率高,和以往培养基中含TPA的培养方法相比,所培养黑素细胞形状变化明显,生长减慢,用于临床移植时更加安全。

AIM:To explore using two step methods to culture melanocytes in order to meet the request of clinical transplantation, eliminate the tumor dangerous of TPA,and increase the reliability of raising cells. METHODS:Melanocytes were cultivated by two steps methods: purification melanocytes were cultivated in culture medium with TPA at first.Then they were subpassaged and cultured in the medium without TPA.The morphology,growth characteristics and cell cycles of melanocytes were investigated in order to observe the character of cultured cells with and without TPA. The cells were transplanted to nude mice to test their oncogenic. RESULT:The purification melanocytes were easily obtained with TPA culture medium;medium without TPA was used when it went down to posterity. The growth speed of melanocyte decreased slightly,the cell cycle proliferated slowly, and their slender dendrites became shortened and broad some cells appear bipolar to polygonal.The experiment of nude mice transplanted with these cells appeared negative. CONCLUSION:The succeed rate of cultivation with two steps method is high. The morphology changes of melanocytes obviously with decreased growth, which suggests that these melanocytes get rid of influence of TPA and are safer to be transplanted in clinic.