Projekte

1) Studying biomineralization in lack of function mouse models

The initiation of biomineralization in vivo is poorly understood but it is speculated that laying down a mineralization competent extracellular matrix enriched in annexins and collagen X is crucial for the initiation of mineralization in cartilage and vasculature. To clarify the relevance of these interactions in biomineralization we have developed transgenic mouse models for these essential factors. Histological and functional analysis of single and multiple knock outs is on their way. Computer tomography imaging systems will help us to quantify the grade of biomineralization in vivo during skeletal development whereas flow cytometry analysis (FACS) enables us to study the calcium homeostasis on cellular level of primary chondrocytes. Whole genome expression profiling (microarray, realtime) of the growth plate was used to identify affected cellular pathways and processes in these mutant mice. We could reveal new factors involved in biomineralization and currently characterizing their influence on biomineralization processes in ex vivo (siRNA knock downs in primary chondrocytes) and in vivo knock down systems (fish, mouse).

2) Using mesenchymal stem cell systems of cartilage and vasculature origin to analyze their differentiation and biomineralization capabilities

Differentiation from mesenchymal stem cells to a calcifying cell represents the important step in endochondral ossification and possibly in calcification of the vasculature. Our studies indicated that annexin A5, one of the most prominent candidates involved in calcification, is found to be expressed in both the vascular and skeletal system (A, C). By combining cell sorting approaches and in vivo reporter gene expression we were able to demonstrate that these annexin A5(+) cells of the vasculature represent vascular associated mesenchymal like stem cells which may resemble skeletal calcification within the vasculature (B). In parallel we are currently collaborating with Dr. Daniele Belluocio (Melbourne, Australia) to combine whole genome expression analysis (microarray) and multiple colour fluorescence activated cell sorting (FACS) to isolate and characterized chondrogenic mesenchymal stem cells of the growth plate. By comparing vascular and chondrogenic stem cells in their differentiation and calcification potential we want to understand essential steps involved in skeletal and vascular calcification to reveal common mechanisms of biomineralization.

A) Expression of annexin A5 - LacZ reporter gene in the vasculature of 11.5 dpc old embryos
B) Isolated annexin A5⁽⁺⁾ mesenchymal like stem cell of the vasculature
C) Expression of annexin A5 - LacZ reporter gene in the skeletal elements of 14.5 dpc old embryos