CAJ | 학술논문

目的：探讨地中海贫血（地贫）患者特异诱导式多能干细胞系（ iPSCs）细胞模型的建立及造血分化能力。方法采集HbH病引产胎儿（α地贫）的脐带血、双重杂合子β地贫输血患者（基因型为IVS－Ⅱ－654／CD17）外周血各1例，分离单核细胞后进行短期培养，核转染再程序化为iPSCs，检测iPSCs未分化状态和多能性标志物。 iPSCs与OP9细胞共培养向造血干细胞诱导分化，进行吉姆萨染色检查和流式细胞术检测造血干细胞的分化结果。结果地贫患者iPSCs表达干细胞多能性标志物Oct4、SSEA4、Tra－1－60、Tra－1－81，碱式磷酸酶染色阳性，悬浮培养可以形成类球形拟胚体，畸胎瘤形成实验显示地贫患者iPSCs可分化为内、中、外胚层细胞。流式细胞术检测结果显示CD34＋细胞分别为8.71％（α－iPSCs分化结果）和4.46％（β－iPSCs分化结果）。结论α、β地贫患者外周血与脐带血可再程序化为 iPSCs，该细胞系具有多能分化特性，与OP9共培养能分化为造血干细胞。
목적：탐토지중해빈혈（지빈）환자특이유도식다능간세포계（ iPSCs）세포모형적건립급조혈분화능력。방법채집HbH병인산태인（α지빈）적제대혈、쌍중잡합자β지빈수혈환자（기인형위IVS－Ⅱ－654／CD17）외주혈각1례，분리단핵세포후진행단기배양，핵전염재정서화위iPSCs，검측iPSCs미분화상태화다능성표지물。 iPSCs여OP9세포공배양향조혈간세포유도분화，진행길모살염색검사화류식세포술검측조혈간세포적분화결과。결과지빈환자iPSCs표체간세포다능성표지물Oct4、SSEA4、Tra－1－60、Tra－1－81，감식린산매염색양성，현부배양가이형성류구형의배체，기태류형성실험현시지빈환자iPSCs가분화위내、중、외배층세포。류식세포술검측결과현시CD34＋세포분별위8.71％（α－iPSCs분화결과）화4.46％（β－iPSCs분화결과）。결론α、β지빈환자외주혈여제대혈가재정서화위 iPSCs，해세포계구유다능분화특성，여OP9공배양능분화위조혈간세포。
Objective To investigate the construction and hematopoietic differentiation of induced pluripotent stem cells ( iPSCs) from thalassemia patients .Methods Cord blood from artificial labored fetus with HbH disease (α-thalassemia) and peripheral blood from a case of heterozygote of β-thalassemia (genotype IVS-II-654/CD17) were obtained.The iPSCs were re-programed from mononuclear cells separated from obtained cord blood and peripheral blood . Hematopoietic stem cell differentiation was induced by OP 9 cell co-culture.The pluripotent markers of iPSCs were detec-ted by immunofluorescence and alkaline phosphatase staining , while embryoid bodies ( EBs) and teratoma forming experi-ments were used to detect the pluripotency .Results Pluripotent biomarkers , including Oct4, SSEA4, tra-1-60 and tra-1-81, and alkaline phosphatase , were positive on iPSCs.EBs were formed by suspended culture .Three germ layer cells were found in the teratoma test .The results of flow cytometry showed that CD 34+cells was 4.46%and 8.71%from cord blood and peripheral sources , respectively, after co-cultured with OP9 cells.Conclusion Cord blood and periph-eral blood MNCs of patients with αorβthalassemia, can be reprogrammed into iPSCs , and differentiated into hematopoi-etic stem cells.