Evaluating the Effect of Cell Culture on Gene Expression in Primary Tissue Samples Using Microfluidic-Based Single Cell Transcriptional Analysis.
Significant transcriptional heterogeneity is an inherent property of complex tissues such as tumors and healing wounds. Traditional methods of high-throughput analysis rely on pooling gene expression data from hundreds of thousands of cells and reporting a population-wide average that is unable to capture differences within distinct cell subsets. Recent advances in microfluidic technology have permitted the development of large-scale single cell analytic methods that overcome this limitation. The increased granularity afforded by such approaches allows us to answer the critical question of whether expansion in cell culture significantly alters the transcriptional characteristics of cells isolated from primary tissue. Here we examine an established population of human adipose-derived stem cells (ASCs) using a novel, microfluidic-based method for high-throughput transcriptional interrogation, coupled with advanced bioinformatic analysis, to evaluate the dynamics of single cell gene expression among primary, passage 0, and passage 1 stem cells. We find significant differences in the transcriptional profiles of cells from each group, as well as a considerable shift in subpopulation dynamics as those subgroups better able to adhere and proliferate under these culture conditions gradually emerge as dominant. Taken together, these findings reinforce the importance of using primary or very early passage cells in future studies.
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. januszyk@stanford.edu.
Michael
M
Januszyk
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. rrennert@stanford.edu.
Robert C
RC
Rennert
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. msorkin@med.umich.edu.
Michael
M
Sorkin
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. zmaan@stanford.edu.
Zeshaan N
ZN
Maan
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. Lisakanata@gmail.com.
Lisa K
LK
Wong
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. arnethaw@stanford.edu.
Alexander J
AJ
Whittam
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. arnethaw@stanford.edu.
Arnetha
A
Whitmore
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. dominikduscher@me.com.
Dominik
D
Duscher
Department of Surgery, Division of Plastic and Reconstructive Surgery, 257 Campus Drive West, Hagey Building GK-201, Stanford, CA 94305-5148, USA. ggurtner@stanford.edu.
Geoffrey C
GC
Gurtner
bioinformatics; cell culture; gene expression; microfluidics; singe-cell; subpopulations; transcriptional analysis