Dynamic Fabrication of Tissue-Engineered Bone Substitutes Based on Derived Cancellous Bone Scaffold in a Spinner Flask Bioreactor System

Abstract

The in vitro dynamic fabrications of tissue-engineered bones were performed to assess the advantages of human adipose-derived stem cells (hADSCs) combined with acellular cancellous bone scaffold coming from fresh pig femur in a spinner flask compared with traditional static culture. In this study, the bio-derived cancellous bone was regarded as a biomimetic scaffold, and its surface appearance was observed under scanning electron microscopy (SEM). Moreover, its modulus of elasticity and chemical composition were measured with universal testing machine (UTM) and infrared detector, respectively. hADSCs were inoculated into cancellous bone scaffold at a density of 1 × 10(6) cells/mL and cultured in spinner flask and T-flask with osteogenic medium (OM) for 2 weeks, respectively. Following to this, the osteogenic differentiation was qualitatively and quantitatively detected with alkaline phosphatase (ALP) kits, and the cell growth and viability were assayed using Live/Dead staining; cell adhesion and extracellular matrix secretion were observed under a SEM. The average pore size of cancellous bone scaffold was 284.5 ± 83.62 μm, the elasticity modulus was 41.27 ± 15.63 MPa, and it also showed excellent biocompatibility. The hADSCs with multidifferentiation potentials were well proliferated, could grow to 90 % fusion within 5 days, and were therefore suitable to use as seed cells in the construction of tissue-engineered bones. After 2 weeks of fabrication, cells were well-distributed on scaffolds, and these scaffolds still remained intact. Compared to static environment, the ALP expression, cell distribution, and extracellular matrix secretion on cancellous bones in spinner flask were much better. It confirmed that three-dimensional dynamic culture in spinner flask promoted ADSC osteogenic differentiation, proliferation, and matrix secretion significantly to make for the fabrication of engineered bone substitutes.

Development of three-dimensional tissue engineered bone-oral mucosal composite models

“…This study demonstrated that improving the quality of culture environment by using spinner bioreactor provided a dynamic microenvironment within the interconnected pores which enhanced nutrient necessary for cells cultivation. This finding is in agreement with many previous studies showing that spinner flask could mitigate the mass transport limitation and promote cells proliferation, differentiation, and expression of osteogenic markers [ 22 , 23 ]. In addition, the cellular trend observed in ROS/HA-TCP construct was in some way consistent with normal osteoblasts’ growth sequence which consists of three principle periods.…”

Beta-tricalcium phosphate granules improve osteogenesis in vitro and establish innovative osteo-regenerators for bone tissue engineering in vivo

“…However, the scaffold is not only a passive support but also provides cells and signals that induce the development of new bone 17 . A wide range of scaffolds that are effective in tissue-engineered bone regeneration have been designed 18 19 20 21 22 23 . However, the scaffolds used in research and clinical applications are suboptimal, and improved scaffolds for in vivo bioreactors are needed.…”

From the macroscale to nanostructures: can tissue engineering recreate bone features?

“…Seeded into a decellularized bone-derived scaffold, adipose tissue-derived stem cells are able to adhere, migrate, grow and differentiate into osteoblastic cells expressing ALP. In these 3D hydrodynamic culture conditions, the level of differentiation was higher than in static conditions [128,129]. However, Rat BM-derived MSCs better differentiated when maintained in a perfused system, rather than in spinner flasks [130].…”

“…De-cellularized / de-mineralized trabecular or cortical bones can be used as biomaterials, as well. They present the advantage to recapitulate the natural ECM features, and allow, therefore, attachment, proliferation and further differentiation of adipose-derived stem cells [129]. Usually, acellular ECM can be obtained after physical, chemical or enzymatic treatments [223,224].…”