Non-technical: This award by the Biomaterials program in the Division of Materials Research to Northwestern University is to study the deposition of single crystalline elements, spicules, made from calcite, a form of calcium carbonate. Many different organisms, from bacteria to humans, use sophisticated organic/inorganic composite materials for mechanical support (e.g., bone), feeding (tooth enamel), defense, and sensing of light, gravity, and the magnetic field. Work in the researcher's laboratory revolves around developing a biotechnological alternative to material synthesis, i.e. modifying an existing biological system rather than re-creating it in the laboratory. Earlier studies by this researcher have demonstrated that the shape and crystallographic growth direction of spicules deposited by primary mesenchyme cells (PMCs) could be controlled using a recombinant protein, vascular endothelial growth factor (VEGF). Building on these results, the team aims to increase the understanding of, and mastery over, the crystal growth process in PMC culture. For this purpose, the effect of VEGF concentration on the structure and composition of the spicule at length scales from nanometers to hundreds of microns will be determined, using sophisticated characterization techniques such as atom probe tomography, tip-enhanced Raman spectroscopy, and X-ray microscopy. Special attention will be given to the distribution of proteins in the biomineral and the surrounding vesicle, using a combination of proteomics and specific antibody-based imaging. Possible functional roles of proteins on nucleation and growth will be established in in vitro assays. In this manner, the team hopes to learn how biological regulation and processing of a complex composite material are connected, essentially linking the biological and the materials genome. This knowledge could impact a wide range of areas from programmed synthesis of bone grafts to large-scale carbon dioxide sequestration. Research findings will be disseminated not only to technical audiences through publications and at conferences, but also through integration in undergraduate laboratory exercises and by engaging undergraduate students in the research as paid laboratory assistants. Finally, members of the team will reach out to local schools and perform experiments with high and middle school students using a mobile laboratory.
Mineralized tissues are sophisticated hybrid materials with highly hierarchical architecture and remarkable control over crystal growth at multiple length scales. Despite great recent progress in bio-inspired material synthesis, many of the hallmarks of biological crystal growth have yet to be reproduced in vitro: polymorph control, curving and/or branching single crystals, and nm scale control of organic-inorganic composites. Clearly, much could be gained by developing a biotechnological alternative to materials synthesis. This researcher's team earlier has developed an in vitro culture system of sea urchin embryo primary mesenchyme cells (PMCs) to control the deposition of single crystalline spicules made from calcite (CaCO3). A fundamental discovery of the prior funding period was that vascular endothelial growth factor (VEGF) signaling controls the shape and crystallographic growth direction of spicules deposited by PMC; the protein does so without directly interacting with the mineral. This provides the unique opportunity to investigate the mechanism underlying this remarkable example of biological control over crystal growth in a well-characterized in vitro cell culture system. The team will do so at three different length scales. At the sub-micron scale, the team will investigate, using a combination of atom probe tomography, tip-enhanced Raman spectroscopy, and X-ray microscopy, the role of organic-mineral interfaces in crystal growth and the impact of the VEGF concentration on the micro/nanostructure and composition of spicules. At the submicron to subcellular length scales, the team will use a combination of quantitative proteomics and imaging using specific antibodies to elucidate the distribution of spicule matrix proteins. Possible functional roles of proteins on nucleation and growth will be established in in vitro assays. Finally, at the system scale, the impact of temporal patterns of VEGF signaling on the crystal branching will be determined. The proposed research will impact the understanding how skeletal elements arise from biological processes at different length scales and how biological processing affects crystal growth. This is a first step to connect developmental biology to the materials genome. This knowledge could impact a wide range of areas from programmed synthesis of bone grafts to large-scale carbon dioxide sequestration. An important component of the proposed activity is an education and outreach-plan that complements the research objectives and is well integrated; its elements are dissemination of research results and methods in the undergraduate curriculum, engagement of students in the research activities, and outreach to local middle and high schools using a mobile laboratory.
