CAJ | 학술논문

目的将多药耐药基因(mdrl)通过逆转录病毒载体导人人胎盘源性间充质干细胞(PMSCs),评价耐药基因导人对P-MSCs干细胞特性的影响.方法采用胰酶消化法自人足月胎盘组织中分离培养间充质干细胞(MSCs),将含有mdr1基因和绿色荧光蛋白(CFP)报告基因的重组逆转录病毒载体转染P-MSCs;应用流式细胞术检测转染后P-MSCs表型特征,四唑盐比色法和碘化丙啶标记测定其增殖活性和细胞周期,透射电镜下观察转染P-MSCs超微结构变化,条件培养液诱导法分析转染后P-MSCs向脂肪细胞、成骨细胞定向分化能力.结果转染P-MSCs表达干细胞表面标志,如CD29,CD44和CD73,细胞倍增时间为23.9 h,生长周期仍符合干细胞增殖的特点,超微结构观察转染P-MSCs表面微绒毛增加,细胞内线粒体丰富,粗面内质网轻度扩张,且在特异诱导条件下具有向成脂、成骨定向分化潜能.结论 mdr1基因体外转染人P-MSCs对其干细胞特性无显著影响,这可为P-MSCs在大剂量肿瘤化疗中的应用提供实验参考.
목적 장다약내약기인(mdrl)통과역전록병독재체도인인태반원성간충질간세포(PMSCs),평개내약기인도인대P-MSCs간세포특성적영향.방법 채용이매소화법자인족월태반조직중분리배양간충질간세포(MSCs),장함유mdr1기인화록색형광단백(CFP)보고기인적중조역전록병독재체전염P-MSCs;응용류식세포술검측전염후P-MSCs표형특정,사서염비색법화전화병정표기측정기증식활성화세포주기,투사전경하관찰전염P-MSCs초미결구변화,조건배양액유도법분석전염후P-MSCs향지방세포、성골세포정향분화능력.결과 전염P-MSCs표체간세포표면표지,여CD29,CD44화CD73,세포배증시간위23.9 h,생장주기잉부합간세포증식적특점,초미결구관찰전염P-MSCs표면미융모증가,세포내선립체봉부,조면내질망경도확장,차재특이유도조건하구유향성지、성골정향분화잠능.결론 mdr1기인체외전염인P-MSCs대기간세포특성무현저영향,저가위P-MSCs재대제량종류화료중적응용제공실험삼고.
Objective To transfer multidrug resistance gene (mdr1) into human placental mesenchymal stem cells (P-MSCs) by retroviral vector and assess the effects of mdr1 gene transduction upon biological features of P-MSCs.Methods Human P-MSCs were isolated from trypsin-digested term placentas and then transduced by reconstructed retroviral vector containing mdr1 gene and green fluorescent protein (GFP) reporter gene.Flow cytometric analysis was employed to determine the immunophenotypes of transfected P-MSCs.And the proliferation and cell cycle were detected by methyl thiazolyl tetrazolium and propidium iodide staining.Uhrastructures of transfected P-MSCs were observed and different induction conditions used to direct the cells to differentiate into adipocytes and osteoblasts.Results The transfected PMSCs still expressed stem cell markers such as CD29,CD44 and CD73.The mean cumulative time of population doubling was 23.9 hours.The cellular cycle retained the proliferative characterization of stem cells.Ultrastructural features of transfected P-MSCs included increased surface microvilli,abundant mitochondria and slightly swollen rough endoplasmic reticulum.Furthermore these transfected cells demonstrated osteogenic and adipogenic differentiation potentials under appropriate conditions.Conclusion The mdr1 gene transduction by retroviral vector in vitro has no significant effect upon biological characteristics of P-MSCs.It might provide experimental references for the application of P-MSCs in high-dose tumor chemotherapy.