Genomics Research Center, Academia Sinica, 128, Sec 2 Academic Road, Taipei 115, Taiwan.
Biomacromolecules. 2012 Nov 12;13(11):3483-93. doi: 10.1021/bm301074j. Epub 2012 Oct 8.
Multilayers of poly-l-lysine/poly-l-glutamic acid (PLL/PLGA) were constructed by layer-by-layer deposition on an end-tethered cationic PLL brush film serving as an initial layer. Increasing the number of coupling layers increased the thickness and the hydration of the films, and decreased the films' shear modulus and serum adsorption. These films were used to culture primary mouse fetal liver cells. Fetal liver stem/progenitor cells (FLSPCs) were isolated and maintained on the PLGA-terminal PLL/PLGA surfaces, forming colonies with clear boundaries that were partially attached to the surface, with cross-sectional areas of ~500 to ~2500 μm(2) after 2 days culture. Long-term studies showed that the cluster size of colonies slowly expanded and was correlated with the surface properties. For example, on the thicker films with shear modulus, G, less than 5 kPa, FLSPCs cluster size was constrained within a small distribution with less than 4000 μm(2) of projected area, whereas on the thinner films with G > 30 kPa, clusters were expanded and widely distributed, with projected areas over 4000 um(2). Immunostaining studies suggested that clusters with a small size maintained the self-renewal characteristics of stem cells, while the expanded clusters were clearly the results of spontaneous differentiation, exhibiting hepatocyte-like properties. On PLL-terminal t-(PLL/PLGA) films, which are less favorable for stem cell cultures than PLGA-terminal t-(PLL/PLGA) films, the cluster size distribution was also correlated with the film thickness, with more clusters of small size preserved on the thicker films. We observed that a soft, hydrated, serum-free surface could restrict the FLSPC expansion, resulting in self-maintenance of FLSPC colonies.
多层层状聚-l-赖氨酸/聚-l-谷氨酸（PLL/PLGA）通过层层自组装在作为初始层的末端接枝阳离子 PLL 刷薄膜上构建。增加偶联层数增加了薄膜的厚度和水合作用，降低了薄膜的剪切模量和血清吸附。这些薄膜用于培养原代小鼠胎肝细胞。胎肝干细胞/祖细胞（FLSPCs）在 PLGA 末端 PLL/PLGA 表面上被分离并维持，在表面部分附着的情况下形成具有清晰边界的集落，在培养 2 天后其横截面积约为 500 至 2500 μm2。长期研究表明，集落的大小缓慢扩大，并与表面特性相关。例如，在剪切模量 G 小于 5 kPa 的较厚薄膜上，FLSPC 集落的大小受到限制，其投影面积小于 4000 μm2；而在 G > 30 kPa 的较薄薄膜上，集落则扩展并广泛分布，投影面积超过 4000 μm2。免疫染色研究表明，具有较小尺寸的集落保持了干细胞的自我更新特性，而扩展的集落显然是自发分化的结果，表现出肝细胞样特性。在 PLL 末端 t-（PLL/PLGA）薄膜上，其对干细胞培养不如 PLGA 末端 t-（PLL/PLGA）薄膜有利，集落大小分布也与薄膜厚度相关，在较厚的薄膜上保留了更多的小尺寸集落。我们观察到，柔软、水合、无血清的表面可以限制 FLSPC 的扩展，从而维持 FLSPC 集落的自我维持。
