期刊
TISSUE ENGINEERING PART C-METHODS
卷 18, 期 4, 页码 302-309出版社
MARY ANN LIEBERT, INC
DOI: 10.1089/ten.tec.2011.0478
关键词
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资金
- Rhode Island Science and Technology Advisory Council
- NIH [R01EB008664-01A1]
The self-sorting of cells into distinct compartments in three-dimensional (3D) microtissues is a process critical to developmental biology, cancer metastasis, and tissue engineering. Although self-sorting has been studied since the 1950s, little quantitative data exist that describe this dynamic process. Here, we describe a recently developed assay designed to quantify the extent and kinetics of self-sorting in 3D. Mixtures of fluorescently labeled normal human fibroblasts (NHF) and hepatocyte (H35) cells were fluorescently labeled, red and green respectively, and seeded onto micro-molded non-adhesive hydrogels. The cells self-assembled into a spheroid and self-sorted with NHFs forming the central core and H35s forming the outer shell. A time course of fluorescent images was used to analyze the ratio of red (NHFs) and green (H35s) fluorescence in concentric hollow cylinders throughout a spheroid and was statistically compared with the fluorescent ratio of the perfectly sorted spheroid. We found that NHFs and H35s, at a 1:1 ratio, sorted to a final extent of 88 +/- 3% at an initial rate of 0.36 +/- 0.06% per minute and reached 50% self-sorted at 2.7 +/- 0.3 h. Studies with varying ratios of NHFs and H35s show that self-sorting and self-assembly are coincident in time when the proportion of NHFs are varied over a 6-fold range (14% to 85%). This method can, thus, be used to characterize the sorting behavior of additional pairs of cells, the effect of drugs, and growth factors that may change the kinetics of the process, and bring an understanding to the cellular mechanisms which control self-sorting.
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