Osteoblasts in a Perfusion Flow Bioreactor—Tissue Engineered Constructs of TiO2 Scaffolds and Cells for Improved Clinical Performance

Abstract

Combining biomaterial scaffolds with cells serves as a promising strategy for engineering critical size defects; however, homogenous cellular growth within large scaffolds is challenging. Mechanical stimuli can enhance bone regeneration by modulating cellular growth and differentiation. Here, we compare dynamic seeding in a perfusion flow bioreactor with static seeding for a synthetic bone scaffold for up to 21 days using the cell line MC3T3-E1 and primary human osteoblast, confocal laser scanning microscopy, and real-time reverse transcriptase-polymerase chain reaction. The secretion of bone-related proteins was quantified using multiplex immunoassays. Dynamic culture improved cellular distribution through the TiO2 scaffold and induced a five-fold increase in cell number after 21 days. The relative mRNA expression of osteopontin of MC3T3-E1 was 40-fold enhanced after 7 and 21 days at a flow rate of 0.08 mL/min, and that of collagen type I alpha I expression was 18-fold after 21 days. A flow rate of 0.16 mL/min was 10-fold less effective. Dynamic culture increased the levels of dickkopf-related protein 1 (60-fold), osteoprotegrin (29-fold), interleukin-6 (23-fold), interleukin-8 (36-fold), monocyte chemoattractant protein 1 (28-fold) and vascular endothelial growth factor (6-fold) in the medium of primary human osteoblasts after 21 days compared to static seeding. The proposed method may have clinical potential for bone tissue engineering.

Three‐dimensional (3D) polylactic acid gradient scaffold to study the behavior of osteosarcoma cells under dynamic conditions

“…The bioreactor has a design similar to that of a bubble column, which enables valuable mixing and perfusion during cell culture experiments, showing significant potential for advancing osteosarcoma applications. Furthermore, recent studies have contributed valuable insights into the role of perfusion in bone tissue engineering 58,59 . These works delve into the specific effects of perfusion on osteoblast behavior within unique scaffold systems, aligning with our efforts to comprehensively address the influence of perfusion on bone formation in our study.…”

“…Furthermore, recent studies have contributed valuable insights into the role of perfusion in bone tissue engineering. 58,59 These works delve into the specific effects of perfusion on osteoblast behavior within unique scaffold systems, aligning with our efforts to comprehensively address the influence of perfusion on bone formation in our study. The comparison between static and dynamic cell cultures revealed differences in cellular proliferation rate (Figure 4).…”

Flourishing tumor organoids: History, emerging technology, and application

“…It simulates microgravity by rotating to create a microgravity, low‐turbulence, low‐shear culture environment for cells, in which cells can grow, self‐aggregate, or aggregate into 3D structures around microcarriers or scaffolds 128 . RWV bioreactors may improve cell distribution and differentiation, together with nutrient and metabolite transport in cell‐inoculated scaffolds 129,130 . Moreover, the adverse effects of shear on cells are reduced, including induction of caspase‐mediated apoptosis, 131 damage to fragile organoid substructures, 132 and perturbed cellular metabolism 133,134 .…”

“…128 RWV bioreactors may improve cell distribution and differentiation, together with nutrient and metabolite transport in cell-inoculated scaffolds. 129,130 Moreover, the adverse effects of shear on cells are reduced, including induction of caspasemediated apoptosis, 131 damage to fragile organoid substructures, 132 and perturbed cellular metabolism. 133,134 RWV is commonly used in bone tissue engineering with good effects on in vivo bone repair in animal studies where the derived tissue constructs resemble natural bone.…”

JANUS: an open-source 3D printable perfusion bioreactor and numerical model-based design strategy for tissue engineering

“…These decisions aim to obtain cell culture conditions that promote a particular cell line's high proliferation and differentiation rates [e.g., mesenchymal stem/stromal cells (Alvarez-Barreto et al, 2011;Bianconi et al, 2023), embryonic stem cells (Marolt et al, 2012), induced pluripotent stem cells (de Peppo et al, 2013)]. Perfusion technology was selected due to its natural double role in bioreactor systems allowing the renovation of the culture medium and, at the same time, applying fluid flow-induced wall shear stress stimuli to the tissue constructs, which has been shown to enhance MSC-mediated bone formation in vitro Wittkowske et al (2016); Schröder et al (2022); Yamada et al (2022). For each microenvironment, finite element models were used to predict the volumetric distributions of fluid flow-induced shear stress and electric field magnitude in culture regions.…”