Itokazu Yutaka, Kitada Masaaki, Dezawa Mari, Mizoguchi Akira, Matsumoto Naoya, Shimizu Akira, Ide Chizuka
Department of Anatomy and Neurobiology, Kyoto University, Graduate School of Medicine, Kyoto, Japan.
Glia. 2006 Jan 1;53(1):32-42. doi: 10.1002/glia.20255.
We previously demonstrated that choroid plexus epithelial (modified ependymal) cells (CPECs) differentiated into astrocytes after grafting into the spinal cord. In the present study, we examined whether CPECs from rats at postnatal 1 day (P1), 7 day (P7), and 8 weeks (P8W) can function as neural progenitor cells that give rise to neurons and glial cells. Cell spheres were produced in cultures of whole tissue of the choroid plexus from the fourth ventricle of rats at each postnatal period. beta-tubulin class III (Tuj-1), glial fibrillary acid protein (GFAP)-, and O4-positive cells differentiated from cell spheres in the differentiation medium. We produced a monoclonal antibody 3E6 specifically labeling microvilli of CPECs. Using this monoclonal antibody, CPECs were isolated from the choroid plexus of P8W rats by cell sorter (FACS). Immunocytochemistry confirmed that there was no contamination from fibroblasts, endothelial cells, macrophages, or Schwann cells in the FACS-isolated 3E6-labeled cells. Cell spheres formed in the cultures of these 3E6-labeled CPECs. After expansion, these cell spheres gave rise to Tuj-1- (5%), GFAP- (45%), and O4-positive cells (0.16%). The remaining cells (45%) were unlabeled neural or glial markers. Some CPECs of the P8W rat were immunohistochemically stained with lineage-associated markers of Musashi-1 and epidermal growth factor-receptor (EGF-R). In addition, infusion of EGF or fibroblast growth factor-2 (FGF2) into the ventricle increased the number of bromodeoxyuridine (BrdU)-positive cells among CPECs from 0.03% (untreated) to 1.14% (38-fold, EGF) and 1.03% (35-fold, FGF2), respectively. These findings indicate that neural progenitor cells exist among CPECs in the rat.
我们之前证明,脉络丛上皮(改良室管膜)细胞(CPECs)移植到脊髓后可分化为星形胶质细胞。在本研究中,我们检测了出生后1天(P1)、7天(P7)和8周(P8W)大鼠的CPECs是否能作为产生神经元和神经胶质细胞的神经祖细胞发挥作用。在每个出生后时期,从大鼠第四脑室脉络丛的全组织培养物中产生细胞球。β-微管蛋白III类(Tuj-1)、胶质纤维酸性蛋白(GFAP)和O4阳性细胞在分化培养基中从细胞球分化而来。我们制备了一种特异性标记CPECs微绒毛的单克隆抗体3E6。使用这种单克隆抗体,通过细胞分选仪(FACS)从P8W大鼠的脉络丛中分离出CPECs。免疫细胞化学证实,FACS分离的3E6标记细胞中没有成纤维细胞、内皮细胞、巨噬细胞或雪旺细胞的污染。这些3E6标记的CPECs培养物中形成了细胞球。扩增后,这些细胞球产生了Tuj-1阳性细胞(5%)、GFAP阳性细胞(45%)和O4阳性细胞(0.16%)。其余细胞(45%)未标记神经或神经胶质标志物。P8W大鼠的一些CPECs用Musashi-1和表皮生长因子受体(EGF-R)的谱系相关标志物进行免疫组织化学染色。此外,向脑室注入表皮生长因子(EGF)或成纤维细胞生长因子-2(FGF2)可使CPECs中溴脱氧尿苷(BrdU)阳性细胞的数量分别从0.03%(未处理)增加到1.14%(38倍,EGF)和1.03%(35倍,FGF2)。这些发现表明大鼠的CPECs中存在神经祖细胞。