Nanoscale topography promotes the survival and proliferation of human β-cells in vitro

Background and aims: Intense research in recent years has aimed at generating sources of human b-like insulin-producing cells for drug discovery and cell transplantation therapy in diabetes. b-cell expansion in tissue culture represents the most obvious approach for the generation of insulin producing cells; however, it has been shown to be difficult. Indeed, cultured human b-cells undergo dedifferentiation and do not replicate. Although the architecture and physical interactions within and surrounding islets are complex and not completely understood, there is significant evidence that islets are heavily influenced by cell–ECM interactions. Aim of the proposed research was to evaluate the suitability of nanostructured scaffolds to support long term culture of human islets of Langerhans and β-cells in vitro. Materials and methods: We used metal oxide layers with tailored nanoscale roughness to fabricate scaffolds for human islet cultures in vitro. We then investigated the suitability of these scaffolds to support long term culture of human islets, through assessment of β-cell differentiation, survival and function in vitro. Finally, using a proteomic approach we evaluate the molecular mechanisms involved. Results: We found that nanostructured substrates significantly increased the number of β-cells and insulin content relative to control (13±2% increased insulin content; p<0.05; 4 different islets preparations ), in long term cultures (up to 25 days). The increased survival of β-cells on nanostructured substrates was due to decreased apoptosis and necrosis, compared to control substrates (15±2% and 20±2% decrease, respectively. P<0.05; 4 different islets preparations). Interestingly, some proliferation was also detected (ki67 positive cells), thus suggesting that these substrates also allow β-cell replication in vitro. Furthermore, only islet β-cells grown over nanostructured scaffolds preserve intact cell morphology with several dispersed insulin granules and the glucose-sensitive insulin secretion in long term cultures. Proteomic analysis confirmed activation of a proliferative program. The process was promoted by a mechanotransductive signalling driven by nanostructured topology via remodelling of the actin cytoskeleton and nuclear architecture. Conclusions: understanding how to manipulate human β-cell survival and proliferation ex vivo may lead to the identification of new biotechnological or pharmacological strategies to treat diabetes.