Enhanced Adipogenic Differentiation of Human Dental Pulp Stem Cells in Enzymatically Decellularized Adipose Tissue Solid Foams.

Abstract

Simple SummaryObesity shortens human lifespan and represents one of the most important public health problems causing significant economic and societal consequences worldwide. However, the current development of physiological human 3D adipose tissue models for in vitro research on preclinical personalized medicine is limited and expensive. Here, we designed, produced, and characterized 3D solid foams using a mixture of bovine collagen I and decellularized human adipose tissue to serve as a 3D matrix mimicking in vivo adipose microenvironment for cell culture purposes. Furthermore, we sought to validate its compatibility for the culture of human mesenchymal stem cells isolated from the dental pulp. We demonstrated that 3D solid foams are able to integrate the stem cells from the dental pulp and provide the appropriate cues to differentiate them into mature adipocytes. The results represent an advance of in vitro 3D models using a human extracellular matrix derived material for future personalized stem cell therapies.Engineered 3D human adipose tissue models and the development of physiological human 3D in vitro models to test new therapeutic compounds and advance in the study of pathophysiological mechanisms of disease is still technically challenging and expensive. To reduce costs and develop new technologies to study human adipogenesis and stem cell differentiation in a controlled in vitro system, here we report the design, characterization, and validation of extracellular matrix (ECM)-based materials of decellularized human adipose tissue (hDAT) or bovine collagen-I (bCOL-I) for 3D adipogenic stem cell culture. We aimed at recapitulating the dynamics, composition, and structure of the native ECM to optimize the adipogenic differentiation of human mesenchymal stem cells. hDAT was obtained by a two-enzymatic step decellularization protocol and post-processed by freeze-drying to produce 3D solid foams. These solid foams were employed either as pure hDAT, or combined with bCOL-I in a 3:1 proportion, to recreate a microenvironment compatible with stem cell survival and differentiation. We sought to investigate the effect of the adipogenic inductive extracellular 3D-microenvironment on human multipotent dental pulp stem cells (hDPSCs). We found that solid foams supported hDPSC viability and proliferation. Incubation of hDPSCs with adipogenic medium in hDAT-based solid foams increased the expression of mature adipocyte LPL and c/EBP gene markers as determined by RT-qPCR, with respect to bCOL-I solid foams. Moreover, hDPSC capability to differentiate towards adipocytes was assessed by PPAR-γ immunostaining and Oil-red lipid droplet staining. We found out that both hDAT and mixed 3:1 hDAT-COL-I solid foams could support adipogenesis in 3D-hDPSC stem cell cultures significantly more efficiently than solid foams of bCOL-I, opening the possibility to obtain hDAT-based solid foams with customized properties. The combination of human-derived ECM biomaterials with synthetic proteins can, thus, be envisaged to reduce fabrication costs, thus facilitating the widespread use of autologous stem cells and biomaterials for personalized medicine.