Techniques to study cancer pathology in vitro lack physiological relevance and can be inappropriate anticancer drug-testing platforms. The crosstalk between stroma, immune components and tumour cells affect cancer progression, such as through altered angiogenesis. Yet, these mechanisms within the prostatic tumour microenvironment remain enigmatic. Star-shaped poly(ethylene glycol)-Heparin (sPEG-Hep) hydrogel offers a flexible 3D platform, where endothelial cells can be stimulated with VEGF to form tube-like capillaries, and fibroblasts can remodel the extracellular matrix through MMP-sensitive crosslinks.
In determining angiogenic changes due to the tumour stroma and a key immunological component, VEGF-stimulated HUVECs were co-cultured with patient derived, unmatched cancer-associated (CAF), or normal-prostatic (NPF) fibroblasts. Moreover, the effect of mast-cell secreted tryptase, a serine protease, was included inside the gels. To validate the effects of tryptase, its inhibitor, Nafamostat Mesylate, was also introduced in culture media. After CD31 staining, HUVECs’ tubes were imaged and quantified in 3D. One-way ANOVA distinguished the statistical differences between groups.
Though preliminary results show nonsignificant trends in groups with tryptase and/or nafamostat mesylate, there are clear changes observed in the CAF/NPF co-culture groups. When combined with CAFs, HUVECs develop increased total number of vessels, which exhibit shorter lengths and reduced branching when compared to the tubes found with NPFs. This could be due to the CAFs’ involvement with malignancy and aberrant vascular development.
sPEG-Hep hydrogel is a rigorous model for analysing tube formation in 3D. Although previous work highlights the involvement of tryptase in inflammation and cancer progression, it remains an enigmatic enzyme, particularly with its interaction inside this hydrogel system. While the results with unmatched CAF/NPF co-cultures show clear trends, future experiments should prioritise the use of CAF/NPF matched-pairs derived from the same patient. This model can potentially be used to analyse the response of patient-specific CAFs with anti-angiogenic/anti-cancer drug therapeutics.