The focus of the ERC Advanced Investigator Grant research group of Prof. Dr. Dr. h. c. Werner E. G. Müller, which includes the groups of Prof. Dr. Xiaohong Wang, Prof. Dr. Dr. Heinz C. Schröder and Dr. Matthias Wiens, concerns the following Topics:
From the evolutionary oldest animals up to humans, we are studying the molecular mechanisms underlying biomineralization. We succeeded to show that the formation of all principle biominerals (silica, calcium carbonate and calcium phosphate), including bone hydroxyapatite (HA), proceeds via enzymatic processes.
These discoveries allowed the development of novel strategies in tissue engineering and repair of bone and cartilage. We could show that the mineralisation of human bone is initiated by a carbonic anhydrase (CA IX) mediated calcium carbonate “bioseed” formation. The initially formed amorphous calcium carbonate (ACC) deposits are then converted into amorphous calcium phosphate (ACP, HA precursor) by carbonate-phosphate exchange. Thereby the phosphate is provided, again enzymatically, by hydrolysis of inorganic polyphosphate (polyP) via the alkaline phosphatase (ALP), a key enzyme of osteoblasts. ACC also stimulates bone formation, but only as long as it is present in the amorphous form. The amorphous ACC rapidly transforms into the more stable crystalline polymorphs vaterite, aragonite, and calcite. We could show that ACC can be stabilized by polyP. The resulting material strongly stimulates bone formation both in vitro and in vivo.
Polyphosphates (polyP) consist of numerous, up to 100 or more phosphate residues which are linked together by high-energy phosphoanhydride bonds. Originally found only in bacteria and yeasts, we later identified polyP also in animals, including humans. We found that comparably high amounts of polyP are present in human osteoblasts and platelets, but also extracellularly in the blood plasma. In particular, we discovered that polyP serves as an extracellular energy storage/signaling molecule. We demonstrated that extracellular ATP can be generated from polyP via the combined action of the cell membrane bound ALP and the adenylate kinase (AK) which catalyzes the reaction 2 ADP ↔ AMP + ATP. This makes polyP interesting especially when used for energy-dependent regeneration of bradytrophic tissues like cartilage.
Using a newly developed procedure, we were able to produce amorphous nano- and microparticles from the calcium salt of polyP. These amorphous Ca-polyP nano/microparticles are stable and become morphogenetically active and biodegradable after transformation into a coacervate in the presence of protein. They show a strong stimulating effect on bone HA synthesis. By combining polyP with other hydrogel-forming polymers, it was possible to develop 3D printable hybrid materials curable by the formation of Ca2+ bridges. These scaffolds are morphogenetically active both in vitro and in vivo.
Building upon our results in basic research, we are developing novel materials for medical applications (bone and cartilage implants, artificial blood vessels and corneas), in particular for 3D-printing/bioplotting. The main focus is on morphogenetically active inorganic polymers, especially energy-rich and smart amorphous polyP nano/microparticles with various metal ions, as well as their combination with other, bioinert materials. This research is funded, among others, by three ERC Proof-of-Concept grants (“Si-Bone-PoC”, “MorphoVES-PoC” and “ArthroDUR”; coordinator: W.E.G. Müller). First regeneratively active implant materials have already been tested in animal experiments and shall be introduced into clinical application. These biocompatible and biodegradable materials allow a fast and complete replacement by the body’s own tissue, without the need of (stem) cells or cytokine/growth factor supplementation.
For example, a morphogenetically active bio-ink with amorphous Ca-polyP microparticles and poly-ε-caprolactone (PCL) has been developed for the potential repair of larger bone defects, as well as Mg-polyP microparticles with viscoelastic properties similar to cartilage for potential treatment of osteoarthritis/arthrosis. Even more effective than Ca-polyP in bone healing are the microparticles obtained with the strontium salt of polyP that not only increased the expression of ALP and BMP-2 in osteoblasts and MSC, but also inhibited the expression of sclerostin, a negative regulator of Wnt signaling and inhibitor of bone cell differentiation and mineralization.
Moreover, with Ca-polyP, we are developing novel strategies for stimulating microvascularization/angiogenesis and for wound treatment. For example, topical application of morphogenetically active polyP microparticles in animal experiments was found to improve wound healing in normal and diabetic mice showing delayed wound healing.