Fibroblast-derived 3D matrix differentially regulates the growth and drug-responsiveness of human cancer cells (original) (raw)
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Tissue Engineering Part A, 2013
Maintenance of cancer stem cells (CSCs) is regulated by the tumor microenvironment. Synthetic hydrogels provide the flexibility to design three-dimensional (3D) matrices to isolate and study individual factors in the tumor microenvironment. The objective of this work was to investigate the effect of matrix modulus on tumorsphere formation by breast cancer cells and maintenance of CSCs in an inert microenvironment without the interference of other factors. In that regard, 4T1 mouse breast cancer cells were encapsulated in inert polyethylene glycol diacrylate hydrogels and the effect of matrix modulus on tumorsphere formation and expression of CSC markers was investigated. The gel modulus had a strong effect on tumorsphere formation and the effect was bimodal. Tumorsphere formation and expression of CSC markers peaked after 8 days of culture. At day 8, as the matrix modulus was increased from 2.5 kPa to 5.3, 26.1, and 47.1 kPa, the average tumorsphere size changed from 37 -6 mm to 57 -6, 20 -4, and 12 -2 mm, respectively; cell number density in the gel changed from 0.8 -0.1 · 10 5 cells/mL to 1.7 -0.2 · 10 5 , 0.4 -0.1 · 10 5 , and 0.2 -0.1 · 10 5 cells/mL after initial encapsulation of 0.14 · 10 5 cells/mL; and the expression of CD44 breast CSC marker changed from 17 -4-fold to 38 -9-, 3 -1-, and 2 -1-fold increase compared with the initial level. Similar results were obtained with MCF7 human breast carcinoma cells. Mouse 4T1 and human MCF7 cells encapsulated in the gel with 5.3 kPa modulus formed the largest tumorspheres and highest density of tumorspheres, and had highest expression of breast CSC markers CD44 and ABCG2. The inert polyethylene glycol hydrogel can be used as a model-engineered 3D matrix to study the role of individual factors in the tumor microenvironment on tumorigenesis and maintenance of CSCs without the interference of other factors.
Scientific Reports, 2016
Three-dimensional (3D) cell cultures represent fundamental tools for the comprehension of cellular phenomena both in normal and in pathological conditions. In particular, mechanical and chemical stimuli play a relevant role on cell fate, cancer onset and malignant evolution. Here, we use mechanically-tuned alginate hydrogels to study the role of substrate elasticity on breast adenocarcinoma cell activity. The hydrogel elastic modulus (E) was measured via atomic force microscopy (AFM) and a remarkable range (150-4000 kPa) was obtained. A breast cancer cell line, MCF-7, was seeded within the 3D gels, on standard Petri and alginate-coated dishes (2D controls). Cells showed dramatic morphological differences when cultured in 3D versus 2D, exhibiting a flat shape in both 2D conditions, while maintaining a circular, spheroid-organized (cluster) conformation within the gels, similar to those in vivo. Moreover, we observed a strict correlation between cell viability and substrate elasticity; in particular, the number of MCF-7 cells decreased constantly with increasing hydrogel elasticity. Remarkably, the highest cellular proliferation rate, associated with the formation of cell clusters, occurred at two weeks only in the softest hydrogels (E = 150-200 kPa), highlighting the need to adopt more realistic and a priori defined models for in vitro cancer studies.
Gelatine methacrylamide-based hydrogels: An alternative three-dimensional cancer cell culture system
Acta Biomaterialia, 2014
Modern cancer research requires physiological, three-dimensional (3-D) cell culture platforms, wherein the physical and chemical characteristics of the extracellular matrix (ECM) can be modified. In this study, gelatine methacrylamide (GelMA)-based hydrogels were characterized and established as in vitro and in vivo spheroid-based models for ovarian cancer, reflecting the advanced disease stage of patients, with accumulation of multicellular spheroids in the tumour fluid (ascites). Polymer concentration (2.5-7% w/ v) strongly influenced hydrogel stiffness (0.5 ± 0.2 kPa to 9.0 ± 1.8 kPa) but had little effect on solute diffusion. The diffusion coefficient of 70 kDa fluorescein isothiocyanate (FITC)-labelled dextran in 7% GelMA-based hydrogels was only 2.3 times slower compared to water. Hydrogels of medium concentration (5% w/v GelMA) and stiffness (3.4 kPa) allowed spheroid formation and high proliferation and metabolic rates. The inhibition of matrix metalloproteinases and consequently ECM degradability reduced spheroid formation and proliferation rates. The incorporation of the ECM components laminin-411 and hyaluronic acid further stimulated spheroid growth within GelMA-based hydrogels. The feasibility of pre-cultured GelMA-based hydrogels as spheroid carriers within an ovarian cancer animal model was proven and led to tumour development and metastasis. These tumours were sensitive to treatment with the anti-cancer drug paclitaxel, but not the integrin antagonist ATN-161. While paclitaxel and its combination with ATN-161 resulted in a treatment response of 33-37.8%, ATN-161 alone had no effect on tumour growth and peritoneal spread. The semi-synthetic biomaterial GelMA combines relevant natural cues with tunable properties, providing an alternative, bioengineered 3-D cancer cell culture in in vitro and in vivo model systems.
Microstructures in 3D Biological Gels Affect Cell Proliferation
Tissue Engineering, 2007
Controlling the microscale environment in 3D matrices for tissue engineering applications is a challenging but necessary goal. In this work, the effect of discrete microscale structures (microrods) on cell proliferation was assessed in three dimensional gels. Microrods were fabricated out of SU-8 with dimensions of 100 by 15 by 15 microns (LxHxW) and incorporated into matrigel seeded with fibroblasts. The 3D microrod-Matrigel composite system inhibited proliferation of both primary and cell-line fibroblasts compared to cells seeded in matrigel alone.
Engineering tumors with 3D scaffolds
Nature Methods, 2007
Microenvironmental conditions control tumorigenesis and biomimetic culture systems that allow for in vitro and in vivo tumor modeling may greatly aid studies of cancer cells' dependency on these conditions. We engineered three-dimensional (3D) human tumor models using carcinoma cells in polymeric scaffolds that recreated microenvironmental characteristics representative of tumors in vivo. Strikingly, the angiogenic characteristics of tumor cells were dramatically altered upon 3D culture within this system, and corresponded much more closely to tumors formed in vivo. Cells in this model were also less sensitive to chemotherapy and yielded tumors with enhanced malignant potential. We assessed the broad relevance of these findings with 3D culture of other tumor cell lines in this same model, comparison with standard 3D Matrigel culture and in vivo experiments. This new biomimetic model may provide a broadly applicable 3D culture system to study the effect of microenvironmental conditions on tumor malignancy in vitro and in vivo.
Gelatin-Based Matrices as a Tunable Platform To Study in Vitro and in Vivo 3D Cell Invasion
ACS Applied Bio Materials, 2019
Hydrogels have been used as synthetic mimics of 3D extracellular matrices (ECM) and their physical properties like stiffness, degradability, porosity have been known to influence the behaviour of encapsulated cells. However, to understand the role of individual properties, the Page 1 of 37 ACS Paragon Plus Environment ACS Applied Bio Materials influence of biophysical cues should be decoupled from biochemical ones. In this study, we have used hydrogels as tunable model matrix to develop 3D cell culture platform for studying cell invasion. Inert polyethylene (glycol) diacrylate (PEGDA) and cell adhesive gelatin methacryloyl (GELMA) were blended in varying compositions, followed by UV mediated photo polymerization to obtain hydrogels with varying stiffness, degradation and cell adhesive properties. We developed two hydrogel matrix systems, namely PEGDA-GELMA (containing larger proportion of PEGDA) and GELMA-PEGDA (containing predominantly GELMA) and characterized them for differences in pore size, swelling ratio, storage modulus, degradability and biocompatibility of the matrix. Both hydrogel systems had similar pore dimensions and swelling behaviour, but PEGDA-GELMA was found to be stiffer and non-degradable while GELMA-PEGDA was softer and degradable. Accordingly, MDA-MB-231 breast cancer cells encapsulated in these matrices showed spheroidal morphology in PEGDA-GELMA hydrogels and were more spindle-shaped in GELMA-PEGDA hydrogels, confirming that size and extent of spreading of cells were influenced by type of these hydrogels. The softer GELMA-PEGDA matrices readily allowed invasion of MDA-MB-231 cells in 3D and showed differences in epithelial-mesenchymal transition (EMT) gene expression of these cells. We further demonstrated the invasion and sprouting of endothelial cells using chick aortic arch assay exhibiting the utility of softer matrices to study 3D cell invasion for multiple applications. We also implanted these matrices in mice and showed that soft gelatin based hydrogels allow cell infiltration in vivo. Results from our study highlight the tunability of this matrix system and the role of matrix constitution in influencing cell invasion in a 3D microenvironment.
Life is three dimensional-as in vitro cancer cultures should be
Advances in cancer research, 2014
For many decades, fundamental cancer research has relied on two-dimensional in vitro cell culture models. However, these provide a poor representation of the complex three-dimensional (3D) architecture of living tissues. The more recent 3D culture systems, which range from ridged scaffolds to semiliquid gels, resemble their natural counterparts more closely. The arrangement of the cells in 3D systems allows better cell-cell interaction and facilitates extracellular matrix secretion, with concomitant effects on gene and protein expression and cellular behavior. Many studies have reported differences between 3D and 2D systems as regards responses to therapeutic agents and pivotal cellular processes such as cell differentiation, morphology, and signaling pathways, demonstrating the importance of 3D culturing for various cancer cell lines.
Engineering cancer microenvironments for in vitro 3-D tumor models
Materials Today, 2015
The natural microenvironment of tumors is composed of extracellular matrix (ECM), blood vasculature, and supporting stromal cells. The physical characteristics of ECM as well as the cellular components play a vital role in controlling cancer cell proliferation, apoptosis, metabolism, and differentiation. To mimic the tumor microenvironment outside the human body for drug testing, two-dimensional (2-D) and murine tumor models are routinely used. Although these conventional approaches are employed in preclinical studies, they still present challenges. For example, murine tumor models are expensive and difficult to adopt for routine drug screening. On the other hand, 2-D in vitro models are simple to perform, but they do not recapitulate natural tumor microenvironment, because they do not capture important three-dimensional (3-D) cell-cell, cell-matrix signaling pathways, and multi-cellular heterogeneous components of the tumor microenvironment such as stromal and immune cells. The three-dimensional (3-D) in vitro tumor models aim to closely mimic cancer microenvironments and have emerged as an alternative to routinely used methods for drug screening. Herein, we review recent advances in 3-D tumor model generation and highlight directions for future applications in drug testing.
Carcinoma Cell-Based Extracellular Matrix Modulates Cancer Cell Communication
Scientific and Academic Publishing, 2022
The extracellular matrix (ECM) is a non-cellular dynamic complex forming a 3D spatial network influencing every cell present in the microenvironment. ECM proteins not only provide scaffolding systems for cells, but they also mediate their functions through cell-cell and cell-ECM interactions. Various tissue-mimicking culture systems or 3D tumor models for tumor studies have been advanced using collagen I, Matrigel, ECM and other biomaterials. The ECM secreted by cancer-associated fibroblasts (CAFs) has been studied for its contribution to tumor growth and its significant role in tumor microenvironment, however, ECM deposited by cancer cells has not been studied in detail. This study examined the role of the ECM derived from a TNBC cell line in cancer cell communications. The decellularized ECM of MDA-MB-231 cells has been prepared and used as the cancer ECM (cECM). The cECM hydrogel was prepared and its 3D spatial network was compared to that of collagen I and Matrigel. Cellular activities of MDA-MB-231 cells in cECM, such as proliferation, migration, cell-cell and cell-ECM interactions were compared to collagen I and Matrigel. E-cadherin as a cell-cell interaction marker, and FAK and α3β1 integrin as cell-ECM interaction markers were considered. Their expression by MDA-MB-231 (MM231) cells in different ECMs were subsequently analyzed. Human umbilical vein endothelial cells (HUVECs) were included in this study to observe the role of cECM in their growth since the tumor microenvironment generates many blood vessels. We found that cECM had modulated cell-cell and cell-matrix interactions for the formation of the tumor microenvironment.