Prof. Ralph Müller

Project 1 – Establishing a 3D In Vitro Co-culture of Osteoclasts and Osteoblasts Bone cells such as osteoclasts, and osteoblasts have significant roles in bone remodeling. So far in our models, we have established long-term culture of osteoblasts and osteocytes. However, bone-resorbing cells such as osteoclasts were not incorporated into the system. In this project, human osteoclast precursors will be introduced, and functional and molecular biology assays will be established along with the time-lapsed microCT scans. Furthermore, the interplay of cell-mediated resorption and formation of extracellular matrix will be evaluated with time-lapsed CT scans and molecular biology assays. This project can be tailored to suit a Bachelor’s Thesis, semester Project or research internship. The students will receive hands-on experience in handling primary cells, bioreactors, 3D printing and conducting molecular biology assays. Project 2 – Optimization of Fibrin as A Bioink for 3D Bioprinting Fibrin-based bioink formulations have garnered research interest due to their excellent biological properties. It is widely used in clinics as a hemostatic agent and surgical sealant. Fibrin and fibrinogen-based proteins are known to have diverse roles in different stages of tissue repair such as cell adhesion, inflammation, angiogenesis, and remodeling. The goal of this project is to develop a 3D printable bio-ink formulation of fibrin and mechanical properties will be tailored to support human mesenchymal stem cell differentiation. This project can be tailored to suit a Masters’ Thesis, Semester Project or Research Internship. The students will receive hands-on experience in primary cell cultivation, inkjet printing, extrusion 3D printing and molecular biology assays.

The projects are multidisciplinary, and students from biology, bioengineering and mechanical engineering backgrounds are welcome. The project goals could be tailored to the student’s interests, expertise, and project requirements.

Dr Chris Steffi (e-mail: chris.steffi@hest.ethz.ch), Postdoc, Laboratory for Bone Biomechanics, ETH Zurich

To address this century-old problem, the Laboratory for Bone Biomechanics has established a novel approach termed “spatial mechanomics”. Spatial mechanomics is defined as the integration of spatially-resolved “omics” with in silico models of the mechanical environment to investigate the interactions between local mechanical environments and cellular / molecular responses (Figure 1). This is a highly cross-disciplinary area of research combining in vivo models, imaging, “omics” techniques, mechanical testing and computational modelling / analyses. There are opportunities for student projects within this area of research that can be tailored to suitable candidates. Ideally applicants will be candidates with an interest or experience in musculoskeletal research, “omics” techniques, computational analyses, mechanical engineering, histology and/or image processing.

We are interested in the following: • To develop a molecular-based understanding of bone healing mechanobiology. • To investigate how this mechano-sensitivity is compromised with age.

Please contact Neashan Mathavan via: nmathavan@ethz.ch

Bone defects that do not heal spontaneously despite surgical stabilization and require further surgical intervention are termed critical sized defects (CSD). They pose dramatic consequences to a person’s well-being and life expectancy due to their impact on the skeletal system’s integrity. Bone graft substitutes (BGS) present a treatment approach by enhancing and facilitating the natural healing capacity of the bone tissue using their osteoconductive and osteoinductive potential and filling up the physical space of lost bone for mechanical support. The key property of a BGS in treatment of a large bone defect is osteoinduction. Physiological processes in bone usually happen on different spatial scales, spanning from the whole organ, to tissue, to cellular and even to the molecular level. Therefore, an ideal imaging setup to **unravel the underlying mechanisms of material-induced osteoinduction** would require a **multiscale approach** to connect orang and tissue level processes with the cellular and subcellular scale. Micro-CT can be considered the gold standard in hierarchical 3D imaging of bone structure and function and allows a non-destructive _ex vivo_ and _in vivo_ image acquisition. **Time-lapsed _in vivo_ micro-CT images** can reveal distinct information on the longitudinal development of implant calcification or resorption. However, its resolution is relatively low and certain details can not be detected. _Ex vivo_ micro-CT on the other hand allows scans of higher resolution and therefore complements the _in vivo_ micro-CT well. The 3D-3D registration of high-resolution _ex vivo_ and low-resolution _in vivo_ micro-CT images generates a multiscale micro-CT imaging approach. Advanced image analysis methods can help identify newly formed bone and generally distinguish bone and the BGS. By combined analysis of multiscale micro-CT measurements of murine femur samples with implanted ectopic or orthotopic BGS, critical 3D mineralization behavior in material-induced osteoinduction can be investigated.

Developing a framework (Python) for the successful (semi-automated) registration of _in vivo_ and _ex vivo_ micro-CT images and advanced image analysis methods of the same murine femur micro-CT scans with BGS for the assessment of 3D mineralization behavior in material-induced osteoinduction. Previous knowledge of python programming and advanced image analysis are of advantage. However, the project goals will be tailored to the student’s interests, expertise, and current project requirements.

Sara Lindenmann, doctoral student (e-mail: sara.lindenmann@hest.ethz.ch), ETH Zürich / Laboratory for Bone Biomechanics (https://www.bone.ethz.ch/) For application, please provide your CV and transcripts of B.Sc. and M.Sc.

The indispensable role of the skeletal system in enabling motion and providing structural support to vital organs underscores the imperative for efficacious interventions in the context of substantial bone defects. Notably, Bone Graft Substitutes (BGS) have emerged as a viable therapeutic avenue, exhibiting success in promoting bone formation through their inherent osteoinductive potential. Despite the considerable promise associated with BGSs, the intricate mechanisms governing material-induced osteoinduction remain elusive. Bohner et al. have recently proposed a novel mechanism termed Sustained Local Ionic Homeostatic Imbalance (SLIHI) as a potential instigator of material-induced osteoinduction. The hypothesis posits that localized depletion of calcium and phosphate during the calcification process may lead to SLIHI, thereby modulating inflammation and instigating an osteoinductive response. Calcium signaling is pivotal in diverse biological and cellular processes, with a particular impact on skeleton mineralization. The continuous remodeling of the skeleton, involving bone resorption and new bone deposition, results in fluctuating extracellular calcium levels corresponding to different phases of remodeling. To comprehensively explore SLIHI, it becomes imperative to scrutinize the spatial distribution and concentration of calcium. This investigation can be effectively conducted through quantitative imaging techniques utilizing calcium-sensitive ratiometric probes. Ratiometric imaging is a scientific approach that utilizes specialized probes to assess variations in calcium concentrations outside cells quantitatively. This technique relies on the simultaneous acquisition of signals at two distinct wavelengths, allowing for the calculation of a ratio that is sensitive to changes in extracellular calcium levels. Here, we aim to assess the impact of ion-regulating BGS on extracellular calcium concentrations and the concurrent influence on adjacent immune cells.

This project seeks to implement ratiometric imaging both in controlled laboratory conditions (in vitro) and within living organisms (in vivo) to examine how bone graft substitutes influence extracellular calcium levels and interact with immune cells. The ultimate goal is to enhance our understanding of these processes to optimize osteoinduction. Candidates with prior familiarity with cell culture and microscopy will be preferably considered. The project goals could be tailored to the student`s interests, expertise, and project requirements.

Dr. Stefanie S. Wissmann, Postdoc (email:stefanie.wissmann@hest.ethz.ch), ETH Zürich / Laboratory for Bone Biomechanics (https://www.bone.ethz.ch/) For application, please provide your CV and transcripts of B.Sc. and M.Sc.