Development and optimization of a dynamic bioreactor for cell culture on PLA scaffolds for bone tissue engineering

In recent years, there has been a significant emphasis on exploiting the approach of tissue engineering to gain a comprehensive understanding of bone structure and how it responds to various stimuli and pharmaceutical agents. Specifically, recent advancements have focused on the construction of scaffolds that can mimic the internal trabecular structure of bone and replicate its chemical and physical characteristics. The primary challenge lies in creating scaffolds that are both biocompatible and durable over time, capable of withstanding the forces exerted by the human skeleton. Within the field of tissue engineering, there is also an aspiration to find an alternative method for cellular cultivation that is not static but can replicate the dynamic environment in which the cell exists. Consequently, my thesis is centered on the development and optimization of a suspension bioreactor for cell culture, designed to recreate the optimal conditions for cell adhesion and growth on the scaffolds. In particular, my thesis is part of the OsteoNet project that aims to develop a reliable and sustainable 3D in vitro cell models of healthy and aged bone tissue. The bone under examination that we aimed to reconstruct is the Human Proximal Ulna. Initially, experiments and optimization analyses of the trabecular structure of the PLA scaffold were conducted using Meshmixer and Fusion360 to achieve values close to physiological ones in order to reproduce healthy and pathological (osteoporotic) bone. These values were subsequently validated through morphological characterization using microCT. From a biological perspective, cell adhesion and growth on the scaffolds inside the bioreactor were analyzed by studying the growth of two types of cells: human osteosarcoma cells and human dental pulp stem cells. To assess the actual cell adhesion to the scaffold, a Wide Field Microscope (Olympus) was used for a qualitative evaluation of how the cells positioned themselves and how uniform their distribution was. The results indicated better uniformity and adhesion compared to the static control used for comparison. It is therefore possible to conclude that a dynamic environment resembling trabecular bone has been reconstructed, which can potentially improve upon the existing static conditions. Another important achievement was the ability to perform seeding and culture within the same chamber. In the future, the idea is to incorporate a control system that can automate the process, making the bioreactor more versatile and commercially viable.