Human-induced pluripotent stem cells produced under xeno-free conditions.
Induced pluripotent stem cells (iPSCs) have radically advanced the field of regenerative medicine by making possible the production of patient-specific pluripotent stem cells from adult individuals. While cell differentiation protocols have been successfully developed, and animal models of human disease have proved that these cells have the potential to treat human diseases and conditions produced as a consequence of aging, degeneration, injury, and birth defects, logistical issues still remain unsolved and hamper the possibility of testing these cells in human clinical trials. Among them is the widely spread use of animal products for the generation and culture of iPSCs. We report here a xeno-free iPSC generation system that addresses all the steps of iPSCs production including the isolation and culture of adult skin fibroblasts, and iPSCs generation, expansion, and maintenance. iPSCs generated with a polycistronic lentiviral vector under xeno-free conditions displayed markers of pluripotency and gave rise to embryoid bodies (EBs) displaying indicators of the 3 primary germ layers. Xeno-free iPSCs injected into nude mice produced classic teratomas, and teratoma explants cultured under conditions favoring fibroblastic cells gave rise to cells morphologically indistinguishable from input cells. Protocols here described will facilitate the implementation of new cellular therapies for preclinical and clinical studies, potentially reducing the regulatory burden without compromising the differentiation potential of the cells.
Ross
Pablo Juan
PJ
Department of Animal Science and Physiology, Michigan State University, East Lansing, Michigan 48824, USA.
Suhr
Steven Thomas
ST
Rodriguez
Ramon Maria
RM
Chang
Eun-Ah
EA
Wang
Kai
K
Siripattarapravat
Kannika
K
Ko
Tak
T
Cibelli
Jose Bernardo
JB
D000818:Animals; D000943:Antigens, Differentiation / Q000235:genetics / Q000378:metabolism; D054337:Cell Dedifferentiation / Q000235:genetics; D002454:Cell Differentiation / Q000502:physiology; D003584:Cytological Techniques / Q000379:methods; D058732:Embryoid Bodies / Q000166:cytology / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism; D015870:Gene Expression / Q000235:genetics; D005822:Genetic Vectors / Q000096:biosynthesis / Q000235:genetics; D005855:Germ Layers / Q000473:pathology; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism / Q000637:transplantation; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics; D016086:Lentivirus / Q000235:genetics; D008297:Male; D051379:Mice; D008819:Mice, Nude; D008875:Middle Aged; D051787:Myogenic Regulatory Factor 5 / Q000378:metabolism; D000071317:Nanog Homeobox Protein; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics; D016601:RNA-Binding Proteins / Q000378:metabolism; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000378:metabolism; D055746:Stage-Specific Embryonic Antigens / Q000378:metabolism; D013724:Teratoma / Q000473:pathology; D014161:Transduction, Genetic
D000943:Antigens, Differentiation; D018398:Homeodomain Proteins; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; C000715187:Lin28A protein, human; C489427:MYC protein, human; C497417:MYF5 protein, human; D051787:Myogenic Regulatory Factor 5; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; D016271:Proto-Oncogene Proteins c-myc; D016601:RNA-Binding Proteins; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; D055746:Stage-Specific Embryonic Antigens; C507976:ZFP42 protein, human; C066584:stage-specific embryonic antigen-4