? 제대혈 유래 중간엽 줄기세포의 순수분리 및 동정 성공. ? 제대혈 유래 중간엽 줄기세포의 대량 배양 방법의 개발. ? 제대혈 유래 다분화능 전구/줄기세포의 분리 및 대량 배양 방법 개발. ? 제대혈 유래 중간엽 줄기세포 및 다분화능 전구/줄기세포의 신경세포로의 분화 유도 성공. ? 신경세포 분화 유도 인자 (Retinoic acid, NGF, DMSO, β-mercaptoethanol, Forskolin, N2 supplement 등)를 첨가한 배지를 이용하여 신경세포 분화유도 확인. ? NGF와 RA
? 제대혈 유래 중간엽 줄기세포의 순수분리 및 동정 성공. ? 제대혈 유래 중간엽 줄기세포의 대량 배양 방법의 개발. ? 제대혈 유래 다분화능 전구/줄기세포의 분리 및 대량 배양 방법 개발. ? 제대혈 유래 중간엽 줄기세포 및 다분화능 전구/줄기세포의 신경세포로의 분화 유도 성공. ? 신경세포 분화 유도 인자 (Retinoic acid, NGF, DMSO, β-mercaptoethanol, Forskolin, N2 supplement 등)를 첨가한 배지를 이용하여 신경세포 분화유도 확인. ? NGF와 RA가 첨가된 신경세포 분화유도 조건에서 약 30% 안팎의 세포가 신경세포로 분화 유도됨. ? Forskolin과 N2 supplement 추가로 약 60%정도의 세포가 신경세포로 분화 유도됨. ? 도파민성 신경세포 (dopaminergic neuron), 콜린성 신경세포 (cholinergic neuron), 가바성 신경세포 (GABAergic neuron) 등의 분화유도 및 그 비율 확인. ? 파킨슨병 동물모델 구축 및 제대혈 유래 단핵세포, 분리/배양된 중간엽 줄기세포, 분리/배양된 전구/줄기세포 또는 분화유도된 세포의 파킨슨병 동물모델에 이식 후 미미한 증상 개선 효과 확인함.
Abstract▼
Umbilical cord blood (UCB) is well known to be a rich source of hematopoietic stem cells with practical and ethical advantages, but the presence of other stem cells, such as mesenchymal stem cells (MSCs), in UCB has been disputed and it remains to be validated. In this study, we examined the ability
Umbilical cord blood (UCB) is well known to be a rich source of hematopoietic stem cells with practical and ethical advantages, but the presence of other stem cells, such as mesenchymal stem cells (MSCs), in UCB has been disputed and it remains to be validated. In this study, we examined the ability of cryopreserved UCB harvests to produce cells with characteristics of MSCs. We were able to obtain homogeneous plastic adherent cells from the mononuclear cell fractions of cryopreserved UCB using our culture conditions. These adherent cell populations exhibited fibroblast-like morphology and typical mesenchymal-like immunophenotypes (CD73+, CD105+, CD166+ etc.). These cells presented the self-renewal capacity and the mesenchymal lineage potential to form bone, fat, and cartilage. These results indicate that cryopreserved human UCB fractions can be used as an alternative source of MSCs for experimental and therapeutic applications. Also, we examined the ability of cryopreserved UCB harvests to produce cells with characteristics of multipotent stem/progenitor cells such as neural precursor. We were able to isolate plastic adherent cells from cryopreserved UCB, which exhibited homogeneous fibroblast-like morphology and immunophenotypes and highly proliferative capacity. They were also positive for nestin and neuroD, the proteins predominantly expressed in neural precursors. Under appropriate conditions for 10-14 days, isolated cells expressed three neural phenotypes: neuron, astrocyte, and oligodendrocyte. Those differentiated cells showed neuronal, astroglial, and oligodendroglial morphology and displayed their respective markers: MAP-1B, NF-L, and NSE for neuron, GFAP for astrocyte, and myelin/oligodendrocyte for oligodendrocyte. We also observed that the cells expressed cell type-specific markers such as tyrosine hydroxylase (TH dopaminergic neuron), acetylcholinesterase (AchE cholinergic neuron), and glutamate decarboxylase (GAD GABAergic neuron). These results indicate that cryopreserved UCB has capacity to produce neural precursor cells and can be used as a potent alternative source of neural stem cells for experimental and clinical applications.
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