Three dimensional in vitro model of invasive tumor spheroids within collagen matrices

Tumor spheroids;collagen gels;not listed

Jimenez Valencia, Angela MariaSun,Sean X. ;

Johns Hopkins University

关键词: Tumor spheroids;    collagen gels;    not listed;   

Others  :  https://jscholarship.library.jhu.edu/bitstream/handle/1774.2/60542/JIMENEZVALENCIA-DISSERTATION-2015.pdf?sequence=1&isAllowed=y

瑞士|英语

来源: JOHNS HOPKINS DSpace Repository

【 摘 要 】

Metastasis, which is the dissemination of cancer cells from solid tumors to distant organs, is the main cause of cancer mortality accounting for ~90% of all cancer-related deaths. The mechanism by which cancer cells spread remains largely unknown due to the difficulty in clinically monitoring the metastatic process. An alternative to study this process is in vitro systems such as the spheroid gel invasion assay, which closely recapitulates the in vivo microenvironment of tumors. This assay consists of embedding multicellular spheroids inside three-dimensional (3D) extracellular matrices (ECM) such as collagen, which is a major component of connective tissues. In this assay, cells emanating from spheroids move through 3D matrices allowing for mechanistic studies of the role of both cell-cell and cell-ECM interactions in cancer cell invasion. Currently, spheroids are embedded inside 3D matrices using well-plates, whereby a truly 3D environment is not always ensured and may give rise to inaccurate invasion analyses. If spheroids are situated at the gel-medium interface they migrate on the surface as opposed to invading through the gel giving the false impression of rapid invasion. To improve the latter, I designed and tested a new experimental set-up using single glass cuvettes, which enables me to fully embed the spheroids in the middle of 3D matrices.To study cancer cell invasion, I embedded spheroids comprised of human fibrosarcoma HT1080 cells inside collagen gels and used live cell microscopy to monitor the invasive front of the spheroids. Analysis on the rate of this invasion suggested that cells invaded in a highly persistent manner. To better understand this invasion process and to obtain improved spatial resolution of cells in the middle of 3D matrices, I developed a method to accurately and consistently cryo-section spheroids-embedded collagen matrices. Computational analysis of cryo-sections showed that cells in the center of the spheroid were rounded in morphology, whereas cells at the edge were elongated. To investigate the association of these morphological differences with cell motility, I tracked individual HT1080 cells inside tumor spheroids that revealed that cells at the periphery invaded in a radial direction. Similar invasion profiles were observed for tumor spheroids embedded in collagen matrices of different densities. I further demonstrated that this invasion process requires myosin II-based cell contractility.These results suggest that the invasion profile of tumor spheroids is an orchestrated process that leads to a highly organized, persistent invasion pattern, fundamentally different than homogeneously embedded single cells in 3D collagen matrices which move randomly.