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Correlative Super Resolution Imaging of the Collagen Mineralization Process

Periodic Reporting for period 1 - SUPERMIN (Correlative Super Resolution Imaging of the Collagen Mineralization Process)

Reporting period: 2018-03-01 to 2020-02-29

Bone tissue is an organic-inorganic composite material that provides mechanical support and protection for our bodies. Its impressive mechanical properties arise from the hierarchical organization of the organic collagen matrix that is mineralized with ultrathin, aligned inorganic crystals of carbonated hydroxyapatite. Despite its importance to the human body, still relatively little is understood about the mechanisms by which collagen mineralization occurs and what the respective roles are of the collagen and other non-collagenous proteins (NCPs) in directing this process. This is because the process is complex: different stages occur over multiple length scales, and many different components are involved. So far, studying collagen mineralization has mainly relied on analyses that require sample-altering preparation methods and lack information about the dynamics; or on simplified in vitro systems that do not necessarily represent what happens in the native bone environment. To understand the role of NCPs in collagen mineralization, we need to study their dynamics and structural interactions with the highest possible resolution and in an environment as close as possible to native bone. Here we build a tissue-engineered bone that has the main characteristic of bone. This model system allowed us to monitor the development of the extracellular matrix in the presence of the different proteins. Besides the unprecedented details that this system provides on the role of selected NCPs in collagen mineralization, it will significantly impact how bone defects and mineralization are studied, and open the door to new treatments for related diseases such as osteogenesis imperfecta and Ehlers-Danlos syndrome.
In this project, we managed to optimize the 3D model system. We have created a reliable model system that represents the process of bone formation, starting from human stem cells that develop into osteoblasts and osteocytes, creating a co-culture. Using this system, we were able to indicate when specific proteins are expressed in relation to the osteoblast maturation and mineralized matrix development. We were able to show that the produced collagen is being mineralized under biological control that has a similar chemical signature of young bone. Our results opened up new research questions, dealing with the time points by which the proteins and the collagen interact for the first time. This interaction may have a significant impact on treating genetic bone-related diseases.
This project allowed us to set up a collaboration on 3D cryo-correlative with Carl Zeiss Company. This unique 3D cryo correlative is beyond the state of the art of imaging techniques, with the hope of launching it during 2020. This 3D cryo correlative imaging workflow will be approachable to many researchers from different fields – setting a new bar for volume imaging in a correlative manner.
3D tissue culture