High-throughput microscopy reveals the impact of multifactorial environmental perturbations on colorectal cancer cell growth

Abstract

Background: Colorectal cancer (CRC) mortality is principally due to metastatic disease, with the most frequent organ of metastasis being the liver. Biochemical and mechanical factors residing in the tumor microenvironment are considered to play a pivotal role in metastatic growth and response to therapy. However, it is difficult to study the tumor microenvironment systematically owing to a lack of fully controlled model systems that can be investigated in rigorous detail.

Results: We present a quantitative imaging dataset of CRC cell growth dynamics influenced by in vivo-mimicking conditions. They consist of tumor cells grown in various biochemical and biomechanical microenvironmental contexts. These contexts include varying oxygen and drug concentrations, and growth on conventional stiff plastic, softer matrices, and bioengineered acellular liver extracellular matrix. Growth rate analyses under these conditions were performed via the cell phenotype digitizer (CellPD).

Conclusions: Our data indicate that the growth of highly aggressive HCT116 cells is affected by oxygen, substrate stiffness, and liver extracellular matrix. In addition, hypoxia has a protective effect against oxaliplatin-induced cytotoxicity on plastic and liver extracellular matrix. This expansive dataset of CRC cell growth measurements under in situ relevant environmental perturbations provides insights into critical tumor microenvironment features contributing to metastatic seeding and tumor growth. Such insights are essential to dynamical modeling and understanding the multicellular tumor-stroma dynamics that contribute to metastatic colonization. It also establishes a benchmark dataset for training and testing data-driven dynamical models of cancer cell lines and therapeutic response in a variety of microenvironmental conditions.

Keywords: colorectal cancer; high-content imaging; liver metastasis; tumor microenvironment.

Figure 1:

The impact of oxygen on CRC growth and treatment response to oxaliplatin. A, Caco2, HT29, and HCT116 cells were cultured in 0.1%, 1%, or 21% oxygen concentration. Cell counts were measured at several time points using Operetta high-content screening platform. B, Growth rate of Caco2, HT29, and HCT116 cells in 0.1%, 1%, or 21% oxygen concentration was determined by CellPD. A 2-sided sign test was used to detect instances where all data trended in a single direction. C, Relative growth rate of Caco2, HT29, and HCT116 cells treated with 0, 0.062, 0.185, 0.555, 1.667, 5 μM oxaliplatin. D, Oxaliplatin IC₅₀ changes in 0.1%, 1%, or 21% oxygen environment.

Figure 2:

The influence of stiffness on CRC growth and treatment response to oxaliplatin. A, HT29-H2BGFP and HCT116-H2BGFP cells were cultured on 0.2 or 2 kPa gel (softwell) or plastic (CellCarrier) plates in 1% or 21% oxygen concentration for 72 hours. Cell counts were measured at several time points by Operetta high-content screening platform, and the growth rate was determined by CellPD. B, Relative growth rate of HT29-H2BGFP and HCT116-H2BGFP cells in response to 0.5 or 5 μM oxaliplatin treatment in 1% or 21% oxygen concentration.

Figure 3:

The effect of liver ECM on the growth of CRC cells and treatment response to oxaliplatin. A, Liver ECM discs were sectioned from acellular liver and seeded with HCT116-H2BGFP cells. B, Snapshots of the disc segmentation process: (1) applying STD filtering and median filtering to the well; (2) applying dilation-reconstruction morphological operations, thresholding and drawing the segmented region over the original image; (3) separating the cells into on-disc (green) and off-disc (red) sets on the basis of the cell location. C, HCT116-H2BGFP cells were cultured on liver ECM disc or monolayer under 1% or 21% oxygen concentration for 72 hours. Cell counts were measured at several time points by Operetta HCS platform, and the growth rate was determined by CellPD. Horizontal black line denotes the mean. ^**P < 0.01 ***P < 0.001 D, Relative growth rate of HCT116-H2BGFP cells in response to 0.5 or 5 μM oxaliplatin treatment under 1% or 21% oxygen concentration. *P < 0.05.