Hyaluronan Synthases and Hyaluronan: Role and Function in the Bone Microenvironment

• Applicant: Dr. Veronika Schönitzer
• Subject Area: Orthopaedics, Traumatology, Reconstructive Surgery
• Term: from 2015 to 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 270807211

Hyaluronan (HA) is assumed to have a regulatory role in the bone remodelling process by influencing the behaviour of mesenchymal stem cells (MSCs), osteoblasts and osteoclasts. The hyaluronan synthases (HAS1, HAS2 and HAS3) which are responsible for the formation of HA are expressed in human MSCs (hMSCs). Although HAS are only active when they are located in the plasma membrane and an intracellular storage pool of the HAS is assumed, the mechanisms controlling the intracellular traffic of HAS are hardly investigated. We hypothesize that HAS interact somehow with the cytoskeleton and that their expression, their transport and/or their activity are regulated via mechanotransduction. We already generated immortalized hMSCs (SCP-1) constitutively expressing eGFP-tagged HAS1, HAS2 and HAS3 respectively in their active form (SCP1-HAS1-eGFP, SCP1-HAS2-eGFP and SCP1-HAS3-eGFP). In this project, we aim to study the mechanical properties of these cells by atomic force microscopy. In addition, we will apply shear stress to the cells and analyse expression, localization and activity of the three HAS isoforms as well as the pathways regulating the cytoskeleton arrangements or the intracellular traffic of HAS. The second target of this project is to analyse how various SCP1-HAS-eGFP affect the behaviour of hMSCs, osteoblasts and osteoclasts. For this purpose, we will coculture hMSCs with SCP1-HAS-eGFP or treat them with exogenous HA of varying molecular weight. We will analyse their behaviour of migration, attachment and spreading, their osteogenic differentiation potential as well as their expression levels of RANKL and OPG. Osteoblasts cocultured with SCP1-HAS-eGFP or treated with HA will be studied with respect to their viability, their mineralization potential and their expression levels of RANKL and OPG. The attachment of osteoclast precursors, osteclastogenesis and expression of RANK by osteoclasts will be investigated in the presence of the SCP1-HAS-eGFP or of exogenous HA. In some bone diseases such as osteogenesis imperfecta, multiple myeloma and osteoporosis, the HA content in the bone or HAS expression in the hMSCs are changed. Understanding the role of HA in bone regeneration and the regulatory mechanisms of HAS in the hMSCs will provide therapeutic starting points for an improved fracture healing in patients suffering from one of these bone diseases. In the long-term perspective, this study may be the basis for the development of new adaptive implant materials.

• DFG Programme: Research Grants