Evaluation of Dynamic Cell Processes and Behavior Using Video Bioinformatics Tools

Abstract

Just as body language can reveal a person's state of well-being, dynamic changes in cell behavior and morphology can be used to monitor processes in cultured cells. This chapter discusses how CL-Quant software, a commercially available video bioinformatics tool, can be used to extract quantitative data on:(1) growth/proliferation, (2) cell and colony migration, (3) reactive oxygen species (ROS) production, and (4) neural differentiation. Protocols created using CL-Quant were used to analyze both single cells and colonies. Time-lapse experiments in which different cell types were subjected to various chemical exposures were done using Nikon BioStations. Proliferation rate was measured in human embryonic stem cell colonies by quantifying colony area (pixels) and in single cells by measuring confluency (pixels). Colony and single cell migration were studied by measuring total displacement (distance between the starting and ending points) and total distance traveled by the colonies/cells. To quantify ROS production, cells were pre-loaded with MitoSOX Red™, a mitochondrial ROS (superoxide) indicator, treated with various chemicals, then total intensity of the red fluorescence was measured in each frame. Lastly, neural stem cells were incubated in differentiation medium for 12 days, and time lapse images were collected daily. Differentiation of neural stem cells was quantified using a protocol that detects young neurons. CL-Quant software can be used to evaluate biological processes in living cells, and the protocols developed in this project can be applied to basic research and toxicological studies, or to monitor quality control in culture facilities.

Mitochondrial Stress Response in Neural Stem Cells Exposed to Electronic Cigarettes

“…Toxicants, such as tobacco smoke, have detrimental effects on the health of stem cells (Kee et al., 2010, Lin et al., 2010, Lin et al., 2009, Lin et al., 2015, Talbot, 2008, Talbot and Lin, 2011). Stem cells exposed to tobacco products or nicotine have diminished regenerative potential because they are less proliferative, migratory, and able to differentiate (Greenberg et al., 2017, Ng et al., 2015, Shaito et al., 2017).…”

Video Bioinformatics Analysis of Human Pluripotent Stem Cell Morphology, Quality, and Cellular Dynamics

“…Live‐cell imaging of hESCs is an emerging field with numerous potential applications, 3,24,25 and several approaches have been developed to evaluate time‐lapse data derived from PSC. Others have used live‐cell imaging to characterize bottlenecks to colony formation when single hESCs are plated at low density 26 .…”

Dynamic blebbing: A bottleneck to human embryonic stem cell culture that can be overcome by Laminin-Integrin signaling

“…A Nikon BioStation IM, which combines an incubator, microscope, and cooled CCD camera, was used to collect time-lapse video data ( Weng et al, 2015 ; Talbot et al, 2014 ; Lin et al, 2015 ). To create time-lapse videos, human prostate cancer cells, mEF, mESC, and hESC were plated on 35 mm culture dishes and incubated in a BioStation IM.…”