Osteoblast-osteoclast co-cultures: A systematic review and map of available literature.

Drug research with animal models is expensive, time-consuming and translation to clinical trials is often poor, resulting in a desire to replace, reduce, and refine the use of animal models. One approach to replace and reduce the use of animal models is to use in vitro cell-culture models. To study bone physiology, bone diseases and drugs, many studies have been published using osteoblast-osteoclast co-cultures. The use of osteoblast-osteoclast co-cultures is usually not clearly mentioned in the title and abstract, making it difficult to identify these studies without a systematic search and thorough review. As a result, researchers are all developing their own methods, leading to conceptually similar studies with many methodological differences and, as a consequence, incomparable results. The aim of this study was to systematically review existing osteoblast-osteoclast co-culture studies published up to 6 January 2020, and to give an overview of their methods, predetermined outcome measures (formation and resorption, and ALP and TRAP quantification as surrogate markers for formation and resorption, respectively), and other useful parameters for analysis. Information regarding these outcome measures was extracted and collected in a database, and each study was further evaluated on whether both the osteoblasts and osteoclasts were analyzed using relevant outcome measures. From these studies, additional details on methods, cells and culture conditions were extracted into a second database to allow searching on more characteristics. The two databases presented in this publication provide an unprecedented amount of information on cells, culture conditions and analytical techniques for using and studying osteoblast-osteoclast co-cultures. They allow researchers to identify publications relevant to their specific needs and allow easy validation and comparison with existing literature. Finally, we provide the information and tools necessary for others to use, manipulate and expand the databases for their needs.

Matrix Vesicles: Role in Bone Mineralization and Potential Use as Therapeutics.

Bone is a complex organ maintained by three main cell types: osteoblasts, osteoclasts, and osteocytes. During bone formation, osteoblasts deposit a mineralized organic matrix. Evidence shows that bone cells release extracellular vesicles (EVs): nano-sized bilayer vesicles, which are involved in intercellular communication by delivering their cargoes through protein-ligand interactions or fusion to the plasma membrane of the recipient cell. Osteoblasts shed a subset of EVs known as matrix vesicles (MtVs), which contain phosphatases, calcium, and inorganic phosphate. These vesicles are believed to have a major role in matrix mineralization, and they feature bone-targeting and osteo-inductive properties. Understanding their contribution in bone formation and mineralization could help to target bone pathologies or bone regeneration using novel approaches such as stimulating MtV secretion in vivo, or the administration of in vitro or biomimetically produced MtVs. This review attempts to discuss the role of MtVs in biomineralization and their potential application for bone pathologies and bone regeneration.

Impact of Culture Medium on Cellular Interactions in in vitro Co-culture Systems.

Co-culturing of cells in in vitro tissue models is widely used to study how they interact with each other. These models serve to represent a variety of processes in the human body such as development, homeostasis, regeneration, and disease. The success of a co-culture is dependent on a large number of factors which makes it a complex and ambiguous task. This review article addresses co-culturing challenges regarding the cell culture medium used in these models, in particular concerning medium composition, volume, and exchange. The effect of medium exchange on cells is often an overlooked topic but particularly important when cell communication via soluble factors and extracellular vesicles, the so-called cell secretome (CS) is being studied. Culture medium is regularly exchanged to supply new nutrients and to eliminate waste products produced by the cells. By removing medium, important CSs are also removed. After every medium change, the cells must thus restore their auto- and paracrine communication through these CSs. This review article will also discuss the possibility to integrate biosensors into co-cultures, in particular to provide real-time information regarding media composition. Overall, the manner in which culture medium is currently used will be re-evaluated. Provided examples will be on the subject of bone tissue engineering.

Rapid Photocrosslinking of Silk Hydrogels with High Cell Density and Enhanced Shape Fidelity.

Silk fibroin hydrogels crosslinked through di-tyrosine bonds are clear, elastomeric constructs with immense potential in regenerative medicine applications. In this study, demonstrated is a new visible light-mediated photoredox system for di-tyrosine bond formation in silk fibroin that overcomes major limitations of current conventional enzymatic-based crosslinking. This photomediated system rapidly crosslinks silk fibroin (<1 min), allowing encapsulation of cells at significantly higher cell densities (15 million cells mL-1 ) while retaining high cell viability (>80%). The photocrosslinked silk hydrogels present more stable mechanical properties which do not undergo spontaneous transition to stiff, ß-sheet-rich networks typically seen for enzymatically crosslinked systems. These hydrogels also support long-term culture of human articular chondrocytes, with excellent cartilage tissue formation. This system also facilitates the first demonstration of biofabrication of silk fibroin constructs in the absence of chemical modification of the protein structure or rheological additives. Cell-laden constructs with complex, ordered, graduated architectures, and high resolution (40 µm) are fabricated using the photocrosslinking system, which cannot be achieved using the enzymatic crosslinking system. Taken together, this work demonstrates the immense potential of a new crosslinking approach for fabrication of elastomeric silk hydrogels with applications in biofabrication and tissue regeneration.