Analysis of invasion 3d culture nih imagej software

Increase in cancer cell invasion via ECM remodeling, invadopodia formation and EMT, as well as drug resistance was recapitulated in the TS-PSC co-culture, and appeared to be mediated by cancer cell-PSC interaction via multiple secreted factors, including IL-6, IL-8, IGF-1, EGF, TIMP-1, uPA, PAI-1, and TSP-1. ResultsД¬o-culture condition was optimized for the growth of TSs, activation of PSCs, and their interaction. Inhibitory effect on cancer cell invasion was compared between gemcitabine and paclitaxel at an equitoxic concentration in PANC-1 TSs co-cultured with PSCs. Conditioned media was also analyzed for secreted factors involved in cancer cell-PSC interactions. Effects of PSC co-culture on ECM fiber network, invasive migration of cancer cells, and expression of epithelial-mesenchymal transition (EMT)-related proteins were examined. PANC-1 cells were cultured as tumor spheroids (TSs) using our previously developed minipillar chips, and co-cultured with PSCs, both embedded in collagen gels.

Preclinical in vitro tumor model of invasive phenotype should incorporate three-dimensional (3D) culture of cancer cells and PSCs in extracellular matrix (ECM) for clinical relevance and predictability. PSCs play significant roles in metastatic progression and chemoresistance through cross-talk with cancer cells. However, the protocol is suitable also for other solid cancer cell lines.Pancreatic ductal adenocarcinoma (PDAC) is a stroma-rich carcinoma, and pancreatic stellate cells (PSCs) are a major component of this dense stroma.

The assay platform was demonstrated using human laryngeal primary and metastatic squamous cell carcinoma cell lines. The invasion is daily imaged, measured, and analyzed using ilastik and Fiji ImageJ software. The spheroids grow in a U-shaped ultra-low attachment plate within the HMDM/fibrin matrix and invade through it. It also includes detailed instructions on imaging and analysis. This protocol provided a simple, reproducible, and reliable 3D human tumor-based spheroid invasion assay using the HMDM/fibrin matrix. It has been shown that HMDM induces migration and invasion of carcinoma cells better than MSDM. Taking into consideration the differences between the human tumor microenvironment and animal-derived matrices, a human myoma-derived matrix (HMDM) was developed from benign uterus leiomyoma tissue. Currently, animal-derived matrices, such as mouse sarcoma-derived matrix (MSDM) and rat tail type I collagen, are mainly used in the spheroid invasion assays. Spheroid invasion assay was developed to provide a suitable 3D in vitro method to study cancer cell invasion. Additionally, this interaction could induce or suppress the expression of several pro- and anti-tumorigenic molecules. This interaction affects cell behavior, such as proliferation and invasion, as well as cell morphology. These assays allow cancer cells to interact with the extracellular matrix. To obtain more reliable in vitro results, several three-dimensional (3D) cell culture assays have been introduced.

However, they lack several basic elements that form the tumor microenvironment. Two-dimensional cell culture-based assays are commonly used in in vitro cancer research.

Analysis of invasion 3d culture nih imagej software