EXTRACELLULAR MATRIX PROTEIN LAMININ DRIVES MOTILITY IN GRANULOCYTE-MACROPHAGE PROGENITORS IN SINGLE-CELL CULTURE

single-cell;GMP;granulocyte-macrophage progenitor;Fluidigm;Polaris;gene expression;motility;extracellular matrix;laminin;microfluidics;Biotechnology

Greene, CassandraObom, Kristina ;

Johns Hopkins University

关键词: single-cell;    GMP;    granulocyte-macrophage progenitor;    Fluidigm;    Polaris;    gene expression;    motility;    extracellular matrix;    laminin;    microfluidics;    Biotechnology;   

Others  :  https://jscholarship.library.jhu.edu/bitstream/handle/1774.2/44649/GREENE-THESIS-2017.pdf?sequence=1&isAllowed=y

瑞士|英语

来源: JOHNS HOPKINS DSpace Repository

【 摘 要 】

Single-cell culture is emerging as a powerful tool for transcriptomics with heterogeneous cell populations, but adequate in vitro extracellular matrix (ECM) conditions have yet to be determined for singulated culture of most cell types. In the absence of cell-to-cell signaling present in bulk culture, characterizing the effect of ECM proteins in single-cell culture is crucial to effective experimental design. In order to address this need and expand the capabilities of the Polaris™ system for single-cell culture, varied ECM proteins were applied for the first time to Polaris microfluidic cell culture integrated fluidic circuits (IFC). Granulocyte-macrophage progenitor cells (GMPs) purified from human cord blood were cultured on laminin-521, collagen 1, truncated vitronectin, fibronectin and unmodified PDMS. Laminin significantly increased cell motility relative to the other ECMs. Sequencing the cDNA libraries generated from these cells revealed differential gene expression for each ECM condition. Genes associated with pathways of motility driven by small Rho GTPases Rac, RhoA, and CDC42 were significantly overexpressed in cells cultured on laminin relative to cells cultured on PDMS. CD34 expression was also far overexpressed in the laminin-cultured cells, suggesting that in as little as 24 hours of single-cell culture, ECM may impact cell fate. This work demonstrates for the first time the application of diverse ECMs on Polaris, and exhibits its utility in performing single-cell studies.