Isolation and identification of tissue stem cell for clinical application

尽管组织干细胞有助于组织的再生，并具有再生医学临床应用的治疗潜力，但与ES细胞相比，组织干细胞的使用存在增殖能力低、可移植能力低等问题。本研究还揭示了睫状体来源的组织干细胞具有通过细胞聚集而不是细胞增殖而分化为神经细胞的潜在能力。为了克服这些问题，我们鉴定和分离了一种新的细胞群，白蛋白(ALB)+a-平滑肌肌动蛋白(a-SMA)VEGFR2+细胞，该细胞也表达ES细胞标志物，并发现在PDGF-BB和VEGF的存在下，该细胞可以分化为a-SMA+平滑肌细胞或CD31+血管内皮细胞，在HGF和EGF的作用下，该细胞在体外可以分化为ALB+肝细胞或CK18+胆管细胞。将未受刺激的VEGFR2+细胞接种于部分肝切除小鼠的脾被膜下，在体内可分化为α-SMA+血管内皮细胞或CD31+血管内皮细胞，经HGF+EGF刺激的VEGFR2+细胞在体内可分化为肝细胞或胆管细胞。在临床治疗性细胞移植中，尽管存在致敏的异反应记忆T细胞的排斥反应，但脐带血来源的CD45-VEGFR2+细胞具有作为内皮祖细胞(EPC)的潜在来源，因为我们使用我们的小鼠肢体缺血模型表明CCL2在缺血肌肉组织中高表达。

Although tissue stem cells contribute to regeneration of tissue and has therapeutic potentials for clinical application of regenerative medicine, the use of the cells has some concerns including low proliferating capacity and low transplantable capacity as compared with ES cells. This study also reveals that ciliary body-derived tissue stem cells have an potential ability to differentiate into neural cells due to cell-aggregation without cell-proliferation. To overcome these issues, we identified and isolated a novel cell population, Albumin(ALB)+a-smooth muscle actin(a-SMA)VEGFR2+ cells, which also expressed the ES cell-expressing markers, and we also found out that the cells could differentiate to a-SMA+ smooth muscle cells or CD31+ vascular endothelial cells in the presence of PDGF-BB and VEGF, and could differentiate to ALB+ hepatocytes or CK18+ bile duct cells in the presence of HGF and EGF in vitro. Moreover, we found out that the non-stimulated VEGFR2+ cells could differentiate to α-SMA+ smooth muscle cells or CD31+ vascular endothelial cells, and HGF+EGF-stimulated VEGFR2+ cells could differentiate to hepatocytes or bile duct cells in vivo, when the cells were inoculated into subcapsule of spleens in partial hepatectomized adult mice.Furthermore, we identified and isolated a cord blood-derived cell population, CD45-CD133+CD34+VEGFR2+ cells, which could form colonies and differentiate to CD133-CD31+ vascular endothelial cells or α-SMA+ smooth muscle cells in the presence of VEGF, IGF, FGF-2,and/or PDGF-BB in vitro. In therapeutic cell transplantation for clinical setting, although there is a concern about rejection by sensitized allo-reactive memory T cells, the cord blood-derived CD45-VEGFR2+ cells has a potential source as endothelial progenitor cell (EPC), because we indicated that CCL2 was highly expressed in the ischemic muscular tissues using our murine limb ischemia models.