Long term culture of cells patterned on glass via membrane-tethered oligonucleotides.
Long term culture of cells patterned on glass via membrane-tethered oligonucleotides.
复制标题
通过膜束缚寡核苷酸在玻璃上形成图案的细胞的长期培养。
DOI：
10.1016/j.biomaterials.2012.09.080
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发表时间：
2013
期刊：
影响因子：
14
通讯作者：
H.
中科院分区：
文献类型：
--
作者：
Sakurai;K.;Hoffecker;I.T.;Iwata;H.
基于寡核苷酸的膜插入物可用作系链以控制细胞与图案化表面的附着，而不干扰内部细胞骨架粘附模式。这种控制可以用作研究细胞-细胞相互作用或不同细胞类型的并排共培养而无需分离/分选的手段。虽然存在与天然细胞骨架机制分离的细胞图案化方法的实用性，但维持这种人工诱导的附着状态的后果仍然未被探索。我们提出了一种通过膜系链单链寡核苷酸与结合在光学透明玻璃基底上的互补单链寡核苷酸杂交来进行细胞二维图案化的方法，这使我们能够表征图案化的HEK 293细胞的长期培养。图案化的基板浸入FBS含介质中，允许吸附的粘附血清蛋白，这允许在最初只通过DNA杂交连接的细胞中的自然细胞骨架粘附模式的传播和参与。我们表明，当细胞试图迁移离开其初始附着点时，共存的附着模式会导致膜结合系链和天然细胞骨架粘附机制之间的竞争。这种竞争最终导致细胞从其指定的模式中逃逸，并且细胞骨架迁移力“胜出”于脂质插入物对细胞膜的亲和力。
Oligonucleotide-based membrane inserts can be used as tethers to control attachment of cells to patterned surfaces without interfering with internal cytoskeletal modes of adhesion. Such control can be employed as a means for study of cell–cell interactions or side-by-side co-culture of different cell types without separation/sorting. While there is utility for cell patterning methods decoupled from natural cytoskeletal mechanisms, the consequences of maintaining this artificially induced state of attachment remains unexplored. We present a method for the 2-dimensional patterning of cells via hybridization of membrane-tethered single stranded oligonucleotides to complimentary single stranded oligonucleotides bound to optically transparent glass substrates which allowed us to characterize the long term culture of patterned HEK293 cells. Patterned substrates immersed in FBS-containing media are shown to permit the adsorption of adhesive serum proteins which allowed for the spreading and engagement of natural cytoskeletal adhesion modes in cells initially attached only through DNA hybridization. We show that the coexisting modes of attachment result in competition between membrane-bound tethers and natural cytoskeletal adhesion machinery as cells attempt to migrate away from their initial points of attachment. This competition ends in the escape of cells from their designated patterns and the ‘winning out’ of cytoskeletal migration forces over the affinity of lipid inserts for the cell membrane.