聚醚砜绒毛膜培养L02细胞的初步研究

目的初步观察一种聚醚砜(polyethersulfone,PES)绒毛膜对人肝细胞系L02细胞培养的支持作用,以探讨该种PES膜材料用于生物人工肝的可行性。方法 PES绒毛膜材料利用非对称性冷却成型技术制备而成。将L02细胞以7.0×104的密度接种于该膜上,分别用悬浮细胞计数法、MTT法、激光共聚焦显微镜、扫描电镜和RT-PCR法来观察L02细胞在该材料上的贴壁率、细胞活性、生长特性及AFP和CYP1A1的基因表达,并以无绒毛型平板膜材料和普通培养板做为对照。结果①PES绒毛膜上L02细胞生长良好,在10h内贴壁率由57.7%增高到91.7%,优于平板膜,与培养板相似;细胞活性8d内增长4至5倍,优于平板膜和培养板。②激光共聚焦显微镜下观察L02细胞在绒毛膜上粘附呈多细胞聚集生长,保持活性;扫描电镜证实了该结果,且逐渐融合形成紧密连接,并见部分细胞呈三维立体生长。③培养于PES绒毛膜上的L02细胞AFP和CYP1A1的表达优于平板膜,与培养板一致。结论 PES绒毛膜能够有效的支持L02细胞在其表面粘附、生长、增殖和AFP及CYP1A1基因的表达,可成为生物人工肝支持系统的膜材料。

A preliminary study on polyethersulfone floss membrane supporting the growth of L02 cell

Objective To observe whether polyethersulfone （PES） floss membrane is a suitable support membrane for hepatocyte L02 cultivation in vitro, and a possible scaffold to be used in the bioartificial liver support system. Methods L02 cells were inoculated with a density of 7.0 × 10^4 on PES floss membrane. By using suspended cell counting, MTT, confocal laser microscopy, scanning electron microscopy and RT-PCR methods, adherence ratio, cell viability, growth characteristics, AFP and CYP1A1 gene expression were evaluated and compared among the cells cultured on PES floss membrane, flat-membrane, and commonly used culture plate. Results ① L02 cells grew well on PES floss membrane. After inoculation for 10 hours, adherence ratio increased from 57.7% to 91.7% in L02 cells on PES floss membrane, which was higher than that of the cells cultured on flat membrane, but was similar to that of the cells on commonly used culture plate. Cell viability increased 4 or 5 times after 8 days, which was higher than that of the cells on other surfaces. ②Confocal microscopy showed that L02 cells adhered on PES floss membrane, grew as multi-cell aggregates, gradually connected by tight junctions, and formed 3 dimensional cell aggregates in some cells, which were confirmed by scanning electron microscopy. ③The expression of AFP and CYP1A1 genes in L02 cells growing on PES floss membrane was higher than that in the cells on flat membrane, but was similar to that in the cells on commonly used culture plate. Conclusion The PES floss membrane can effectively support adhesion, growth, proliferation, and expression of AFP and CYP 1A 1 of the L02 cells inoculated on its surface. Therefore, this membrane may be used as a novel scaffold biomaterial for bioartificial liver.