基因工程苯丙氨酸解氨酶对多发性骨髓瘤U266细胞的作用研究  

An in vitro Study of Cytotoxic and Antineoplastic Effect of Gene Engineering Phenylalanine Ammonia-lyase on Multiple Myeloma Cell Iine U266

作　　者：陈星[1] 王璞[2] 刘晋纬[3] 肖白[1] 陈文明[3] 刘敬忠[1] 

机构地区：[1]首都医科大学附属北京朝阳医院基础医学研究中心,100020 [2]首都师范大学生命科学学院,北京100037 [3]首都医科大学附属北京朝阳医院血液与肿瘤科

出　　处：《医学研究杂志》2008年第10期45-47,共3页Journal of Medical Research

摘　　要：目的为寻找对多发性骨髓瘤新的治疗方法，从基因工程乳酸乳球菌提纯苯丙氨酸解氨酶（PAL），探讨PAL酶对多发性骨髓瘤U266细胞系的影响作用。方法利用已构建成功的高表达PAL的基因工程乳酸乳球菌，结合物理、化学、生物学等方法，设计一种简便、高效提纯PAL酶的方法。培养人多发性骨髓瘤U266细胞系，在不同浓度基因工程PAL酶作用后，以MTT法检测细胞增生；流式细胞仪检测细胞凋亡；HPLC检测细胞培养液中苯丙氨酸（phe）浓度。结果①用本研究所设计方法从基因工程乳酸乳球菌NZ3900／pNZ8149-pal^art中提取的PAL酶比活性可达同量原菌的92．5％；②0．1～2．0U／ml的PAL提取液作用24h后能显著抑制U266细胞生长，并呈剂量依赖性；③只添加提取过程所用试剂的对照组细胞生长无明显抑制；④0．1U／ml、0．2U／ml、2．0U／ml的PAL酶作用24h后的MM细胞早期凋亡率分别为（21．92±0．95）％、（50．64±0．86）％、（90．86±2．91）％；⑤PAL酶作用24h后细胞培养液中phe浓度随PAL酶活性增加而降低。结论本研究所设计方法可从基因工程乳酸菌高效提取PAL酶；提取过程中的添加试剂对MM细胞生长无明显抑制作用；PAL酶能显著抑制MM细胞体外生长，诱导细胞早期凋亡；PAL酶可降低培养液中的phe水平，且phe浓度与MM细胞生长的抑制作用具有相关性。Objective To study the effects of phenylalanine ammonia - lyase （PAL） extract from gene engineering Lactococeus Lactis （ NZ3900/pNZ8149 - palart） on proliferation and apoptosis of human multiple myeloma cell line U266. Methods A new method was established with many techniques of physics, chemistry and biology to extract PAL from gene engineering L. L, then the activities and purities of the extraction were detected and analyzed. The proliferation inhibition rate of U266 cells was assayed by MTT analysis. Cell apoptosis were evaluated by flow eytometry （FCM） and phenylalannie （phe） concentration of cell curture were measured by HPLC. Result The PAL enzyme activities of the extraction were 92.5% of the same weight living gene engineering L. L. After U266 cells were treated with PAL extraction （ the concentration from 0. 1 U/ml to 2.0U/ml） for 24h, the cell proliferation was markedly inhibited with a dose related manner. When U266 cells were treated with 0. 1 U/ml,0. 2U/ml,2.0 U/ml PAL for 24h, the early apoptosis rate revealed by Annexin V/PI were 21.92%、50. 64% and 90. 86% respectively, being higher than that of the blank controls （0. 88% and 1. 12%±0. 15% , P 〈 0. 01 ）. The phe concentration of cell curtnre was reduced after treated with PAL. Conclusion The PAL extract from gene engineering L. L could obviously inhibit the proliferation and induce the apoptosis of U266 cells, which maybe attributed to the reduction of phe concentration.

关 键 词：苯丙氨酸解氨酶 基因工程 多发性骨髓瘤 凋亡 

分 类 号：R977.3[医药卫生—药品] R733.3[医药卫生—药学]