Gmyr Valéry, Belaich Sandrine, Muharram Ghaffar, Lukowiak Bruno, Vandewalle Brigitte, Pattou François, Kerr-Conte Julie
Cell Therapy of Diabetes, INSERM ERIT-M 0106, University Hospital of Lille, 1 place de Verdun, Lille 59045, France.
Biochem Biophys Res Commun. 2004 Jul 16;320(1):27-33. doi: 10.1016/j.bbrc.2004.05.125.
Generating human insulin-secreting cells for cell therapy of diabetes represents a highly competitive world challenge. Human ductal cells can give rise to islets in vivo and in vitro. The goal of this study was to devise a rapid sorting method to highly purify human ductal cells from pancreatic tissue using a pan-ductal membrane antibody carbohydrate antigen 19-9 (CA19-9). Human pancreatic sections confirmed antibody specificity. The human exocrine fraction (30% ductal cells) was sorted with magnetic bead technology or by FACS. Immunocytochemistry post-sorting determined ductal cell content. The manual magnetic bead technique resulted in 74%+/-2 (n = 4) CA19 positive cells. Whereas the automated AutoMACS technique (n = 5) yielded 92.6%+/-0.5 CA19-9 positive cells with only a minor beta cell contamination (0.2%+/-0.03); cell yield post-sorting was 12.9%+/-2.5 (1.69+/-0.41 x 10(6) cells) with 51.7%+6.5 (n = 5) viability post-sorting. The FACS (n = 6) resulted in 97.1%+/-0.82 CA19-9 positive cells, a cell yield of 25.5%+/-5.6 (5.03+/-1.0 x 10(6)), with 72.1%+/-6.1 viability post-sorting.
生成用于糖尿病细胞治疗的人胰岛素分泌细胞是一项极具竞争力的全球性挑战。人导管细胞在体内和体外均可产生胰岛。本研究的目的是设计一种快速分选方法，使用全导管膜抗体碳水化合物抗原19-9（CA19-9）从胰腺组织中高度纯化人导管细胞。人胰腺切片证实了抗体的特异性。使用磁珠技术或荧光激活细胞分选术（FACS）对人外分泌部分（30%为导管细胞）进行分选。分选后通过免疫细胞化学测定导管细胞含量。手动磁珠技术得到74%±2（n = 4）的CA19阳性细胞。而自动AutoMACS技术（n = 5）产生92.6%±0.5的CA19-9阳性细胞，仅有少量β细胞污染（0.2%±0.03）；分选后的细胞产量为12.9%±2.5（1.69±0.41×10⁶个细胞），分选后活力为51.7%±6.5（n = 5）。FACS（n = 6）得到97.1%±0.82的CA19-9阳性细胞，细胞产量为25.5%±5.6（5.03±1.0×10⁶），分选后活力为72.1%±6.1。
