Mouse-Induced Pluripotent Stem Cells Generated Under Hypoxic Conditions in the Absence of Viral Infection and Oncogenic Factors and Used for Ischemic Stroke Therapy

Abstract

Induced pluripotent stem (iPS) cells are considered as having the greatest potential for use in cell-based therapies. However, at least two hurdles remain: integrating viral transgenes and introducing the c-Myc and Klf4 oncogenes. In a previous study, fibroblasts were incapable of generating iPS cells in the absence of both oncogenes and viral infection. For the present study, we tested our hypothesis that iPS cells can be generated without oncogenes and viral infection under hypoxic conditions and used for cell therapies. By avoiding oncogenic factors and virus integration, this strategy would decrease the potential for cancer formation. According to our observations, the repeated transfection of two expression plasmids (Oct4 and Sox2) into mouse embryonic fibroblasts (MEFs) and combined hypoxic condition resulted in the generation of a novel iPS cell. At 6 h post-transfection, MEFs were subjected to hypoxic conditions (3% O2) for 24 h; this procedure was repeated four times. The resulting MEFs were seeded on feeder cells on day 9; iPS cell clones were observed 12 days post-seeding and designated as iPS-OSH. Data for cell morphology, stem cell marker staining, gene expression profiles, and embryonic body, teratoma, and chimeric mouse formation indicated iPS-OSH pluripotent capability. Neural precursor cells differentiated from iPS-OSH cells were used to treat an ischemic stroke mouse model; results from a behavior analysis indicate that the therapeutic group surpassed the control group. Further, iPS-OSH-derived neural precursor cells differentiated into neurons and astrocytes in mouse stroke brains. In conclusion, we generated a novel iPS-OSH in the absence of viral infection and oncogenic factors and could use it for ischemic stroke therapy.

Opportunities and Challenges: Stem Cell‐Based Therapy for the Treatment of Ischemic Stroke

“…Recently, a novel strategy was reported that using a combination of Oct4 and Sox2 plasmid transfection with hypoxia conditioning to generate iPS cells (iPS-OSH cells). iPS-OSH-derived NPCs differentiated into neurons and astrocytes after being injected into ischemic mice brain, accompanied by improved neurobehavioral recovery [45].…”

Regulated and Unregulated Clinical Trials of Stem Cell Therapies for Stroke

“…By targeting the subacute and chronic phases of stroke, stem cells significantly extend the effective period of intervention, permitting a larger patient population to benefit from this prospective treatment. From the many cell types which have been transplanted in the laboratory, a small fraction have reached clinical trials for cell therapy in stroke, including fetal cells, NT2N cells, CTX0E3, embryonic stem cells, neural stem/progenitor cells, umbilical cord blood, amnion, adipose, and induced pluripotent stem cells [6], [7], [8], [9], [10]. Bone marrow-derived cells have been distinguished in preclinical studies and ongoing clinical trials largely due to maintaining a consistent safety profile when used in other diseases [11], [12].…”

Recent Advances in Stem Cell-Based Therapeutics for Stroke

“…From the initial study of fetal cell transplantation into stroke animals, varying levels of histological and behavioral recovery have been demonstrated in NT2N, CTX0E3, embryonic stem cells, hematopoietic stem cells, neural stem cells, adult tissue-derived stem cells, and induced pluripotent stem cells [7–11, 22]. The quest for the optimal cell type for transplantation therapy has largely been dictated by ethical and logistical issues [23, 24].…”