![analysis of invasion 3d culture nih imagej software analysis of invasion 3d culture nih imagej software](https://onlinelibrary.wiley.com/cms/asset/02b35caf-87eb-459a-af24-f8ed530664b8/cas14664-fig-0005-m.jpg)
Amongst breast cancer subtypes we found that the stroma at the invasive region of the more aggressive Basal-like and Her2 tumor subtypes was the most heterogeneous and the stiffest when compared to the less aggressive luminal A and B subtypes. The linearization of collagen was visualized as an overall increase in tissue birefringence and was most striking at the invasive front of the tumor where the stiffness of the stroma and cellular mechanosignaling were the highest. Our analysis revealed that human breast cancer transformation is accompanied by an incremental increase in collagen deposition and a progressive linearization and thickening of interstitial collagen. We undertook a biophysical and biochemical assessment of stromal–epithelial interactions in noninvasive, invasive and normal adjacent human breast tissue and in breast cancers of increasingly aggressive subtype. Nevertheless, the relationship between tissue and extracellular matrix stiffness and human breast cancer progression and aggression remains unclear. The 3D-3-culture constitutes a novel tool to study tumour-immune interaction and macrophage plasticity in response to external stimuli, such as chemotherapeutic and immunomodulatory drugs.Tumors are stiff and data suggest that the extracellular matrix stiffening that correlates with experimental mammary malignancy drives tumor invasion and metastasis. In conclusion, the crosstalk between the ECM and tumour, stromal and immune cells in microencapsulated 3D-3-culture promotes the activation of monocytes into TAM, mimicking aggressive tumour stages. Specifically, the macrophage phenotype was modulated upon treatment with the CSF1R inhibitor BLZ945, resulting in a decrease of the M2-like macrophages. The 3D-3-culture was challenged with chemo- and immunotherapeutic agents and the response to therapy was assessed in each cellular component. Importantly, we show that both the monocytic cell line THP-1 and peripheral blood-derived monocytes infiltrate the tumour tissue and transpolarize into an M2-like macrophage phenotype expressing CD68, CD163 and CD206, resembling the TAM phenotype in NSCLC. We demonstrate that the 3D-3-culture recreates an invasive and immunosuppressive TME, with accumulation of cytokines/chemokines (IL4, IL10, IL13, CCL22, CCL24, CXCL1), ECM elements (collagen type I, IV and fibronectin) and matrix metalloproteinases (MMP1/9), supporting cell migration and promoting cell-cell interactions within the alginate microcapsules. In this work, we explored a culture platform based on alginate microencapsulation and stirred culture systems to develop the 3D-3-culture, which entails the co-culture of tumour cell spheroids of non-small cell lung carcinoma (NSCLC), cancer associated fibroblasts (CAF) and monocytes. This particularly holds true for prediction of therapeutic response(s) to standard therapies and interrogation of efficacy of TME-targeting agents. Recapitulation of the interaction between the different cellular players of the TME, along with the extracellular matrix (ECM), is critical for understanding the mechanisms underlying disease progression.
![analysis of invasion 3d culture nih imagej software analysis of invasion 3d culture nih imagej software](https://www.dovepress.com/cr_data/article_fulltext/s249000/249756/img/OTT_A_249756_O_F0003g.jpg)
Most aggressive solid tumours have high levels of myeloid cell infiltration, namely tumour associated macrophages (TAM).
![analysis of invasion 3d culture nih imagej software analysis of invasion 3d culture nih imagej software](https://www.researchgate.net/profile/Nandana-Bhardwaj/publication/305663814/figure/fig8/AS:668713992847366@1536445248330/A-Schematic-representation-of-fabrication-of-hydrogels-and-B-macroscopic-appearance_Q640.jpg)
The tumour microenvironment (TME) shapes disease progression and influences therapeutic response.