Bone Abstracts

Objectives: Strong coordination between osteoblasts and bone marrow cells is fundamental for the regulation of healthy bone turnover and other (patho)physiological processes. Extracellular vesicles (EVs) mediate communication between cells via horizontal transfer of proteins and nucleic acids. Osteoblasts secrete EVs in the form of matrix vesicles involved in bone mineralization, however, information about a role in intercellular communication is still lacking. In this study, we focus on the characterization of human osteoblast-secreted EVs and study their role as mediators of communication with neighbouring cells in their bone marrow microenvironment.

Methods: We used a human pre-osteoblast-based in vitro bone formation model to isolate EVs at various time-points during osteoblast differentiation and mineralization by a series of ultracentrifugation steps. We characterized the EVs by electron and atomic force microscopy and proteomics. Furthermore, we studied their interaction with neighbouring cells by fluorescent labelling and flow cytometric analysis.

Results: Microscopic analyses demonstrated that osteoblast EVs are very heterogenic in size and morphology. Mass spectrometry-based proteomic analyses identified known matrix vesicle proteins (annexins, phosphatases, etc.) and an interesting range of membrane and signalling proteins that may be linked to cell communication. Fluorescently labelled osteoblast EVs were internalized by neighbouring cells such as CD34⁺ hematopoietic stem cells (HSCs) as well as bone-metastasizing prostate cancer cells in a dose-dependent manner. Cultures of CD34⁺ HSCs with osteoblast EVs led to a donor-dependent two to three-fold increased proliferation of the CD34⁺ expressing cells.

Conclusion: We demonstrated that osteoblasts secrete EVs that are taken up by the cells residing in the surrounding microenvironment. Osteoblast EVs are specifically packaged not only with common matrix vesicle proteins showing their importance in bone metabolism but also with proteins not-primarily linked to mineralization suggestive of a novel mechanism of intercellular communication.

This study has been supported by Erasmus Stem Cell Institute.