Generate Induced Pluripotent Stem Cells From Human Peripheral Blood Mononuclear Cell

Objective: Induced pluripotent stem cells(iPSCs)have a lot of advantages.first,it can be generated from various types of cells,second,it also can be btained from the patients themselves directly,and in vitro it can be successfully differentiated into hematopoietic progenitors.In this study,we will discuss how to generate induced pluripotent stem cells from human peripheral blood mononuclear cell,it will lay a foundation of generation of G6 PD deficiency-induced pluripotent stem cells.Methods: Collect healthy people's peripheral venous blood,and isolate peripheral blood mononuclear cell with Ficoll density gradient centrifugation.Then we infect peripheral blood mononuclear cell with sendai virus,which contain four reprogramming factors,Oct4,Sox2,Klf4 and c-Myc.Finally,plate infected cells on feeder(MEF)culture dishes.Identify the PBMC-derived i PSCs by observing of cell morphology,TRA1-60 live staining,and RT-PCR.Results: stem cell like colonies gradually emerged at the day 15 afterinfecting sendai virus into peripheral blood mnonuclear cells.At the day 24,the large round or oval stem cell like clones,with clear border,were obtained.Live staining shows that PBMC-derived iPSCs expressed stem cell specific surface antigen TRA1-60,and the results of RT-PCR also shows the PBMC-derived iPSCs carried stem cell pluripotent related genes-Lin28,Oct4 and Nanog.Conclusion: Induced pluripotent stem cells can be obtained by infecting peripheral blood mononuclear cells with sendai virus,which contain four reprogramming factors,Oct4,Sox2,Klf4 and c-Myc.PBMC-derived iPSCs are similar to embryonic stem cells in morphological characteristics and the expressing of multipotent genes.