The immense diversity of extracellular matrix (ECM) proteins confers distinct biochemical and biophysical properties that influence cell phenotype. The ECM is highly dynamic as it is constantly deposited, remodelled, and degraded during development until maturity to maintain tissue homeostasis. The ECM’s composition and organization are spatiotemporally regulated to control cell behaviour and differentiation, but dysregulation of ECM dynamics leads to the development of diseases such as cancer. The chemical cues presented by the ECM have been appreciated as key drivers for both development and cancer progression. However, the mechanical forces present due to the ECM have been largely ignored but recently recognized to play critical roles in disease progression and malignant cell behaviour. Here, we review the ways in which biophysical forces of the microenvironment influence biochemical regulation and cell phenotype during key stages of human development and cancer progression.

Despite advances in cancer treatment, breast cancer remains the second foremost cause of cancer mortality among women, with a high rate of relapse after initial treatment success. A subpopulation of highly malignant cancer cells, known as cancer stem cells (CSCs), is suspected to be linked to metastasis and relapse. Targeting of CSCs may therefore provide a means of addressing cancer-related mortality. However, due to their low population in vivo and a lack of proper culture platform for their propagation, much of the CSC biology remains unknown. Since maintenance of CSCs is heavily influenced by the tumor microenvironment, this study developed a 3D culture platform that mimics the metastatic tumor extracellular matrix (ECM) to effectively increase CSC population in vitro and allow CSC analysis. Through electrospinning, nanofibers that were aligned, porous, and collagen-coated were fabricated from polycaprolactone to recreate the metastatic tumor ECM assemblage. Breast cancer cells seeded onto the nanofiber scaffolds exhibited gross morphology and cytoskeletal phenotype similar to invasive cancer cells. Moreover, the population of breast cancer stem cells increased in nanofiber scaffolds. Analysis of breast cancer cells grown on the nanofiber scaffolds demonstrated an upregulation of mesenchymal markers and an increase in cell invasiveness suggesting the cells have undergone epithelial-mesenchymal transition. These results indicate that the fabricated nanofiber scaffolds effectively mimicked the tumor microenvironment that maintains the cancer stem cell population, offering a platform to enrich and analyze CSCs in vitro.

Advances in stem cell technologies, revolutionising regenerative therapies and advanced in vitro testing, require novel cell manufacturing pipelines able to cope with scale up and parallelisation. Microdroplet technologies, which have transformed single cell sequencing and other cell-based assays, are attractive in this context, but the inherent soft mechanics of liquid-liquid interfaces is typically thought to be incompatible with the expansion of induced pluripotent stem cells (iPSCs), and their differentiation. In this work, we report the design of protein nanosheets stabilising liquid-liquid interfaces and enabling the adhesion, expansion and retention of stemness by iPSCs. We use microdroplet microfluidic chips to control the formulation of droplets with defined dimensions and size distributions and demonstrate that these sustain high expansion rates, with excellent retention of stem cell marker expression. We further demonstrate that iPSCs cultured in such conditions retain the capacity to differentiate into cardiomyocytes and demonstrate such process on droplets. This work provides clear evidence that local nanoscale mechanics, associated with interfacial viscoelasticity, provides strong cues able to regulate and maintain pluripotency, as well as to support commitment in defined differentiation conditions. Microdroplet technologies appear as attractive candidates to transform cell manufacturing pipelines, bypassing significant hurdles paused by solid substrates and microcarriers.