Rationale: Fetuin-A is a liver-derived plasma protein involved in the regulation of calcified matrix metabolism.Biochemical studies showed that fetuin-A is essential for the formation of protein-mineral complexes, called calciprotein particles (CPPs). CPPs must be cleared from circulation to prevent local deposition and pathological calcification.Objective: We studied CPP clearance in mice and in cell culture to identify the tissues, cells, and receptors involved in the clearance. Methods and Results:

Muscle and bone interact via physical forces and secreted osteokines and myokines. Physical forces are generated through gravity, locomotion, exercise, and external devices. Cells sense mechanical strain via adhesion molecules and translate it into biochemical responses, modulating the basic mechanisms of cellular biology such as lineage commitment, tissue formation, and maturation. This may result in the initiation of bone formation, muscle hypertrophy, and the enhanced production of extracellular matrix constituents, adhesion molecules, and cytoskeletal elements. Bone and muscle mass, resistance to strain, and the stiffness of matrix, cells, and tissues are enhanced, influencing fracture resistance and muscle power. This propagates a dynamic and continuous reciprocity of physicochemical interaction. Secreted growth and differentiation factors are important effectors of mutual interaction. The acute effects of exercise induce the secretion of exosomes with cargo molecules that are capable of mediating the endocrine effects between muscle, bone, and the organism. Long-term changes induce adaptations of the respective tissue secretome that maintain adequate homeostatic conditions. Lessons from unloading, microgravity, and disuse teach us that gratuitous tissue is removed or reorganized while immobility and inflammation trigger muscle and bone marrow fatty infiltration and propagate degenerative diseases such as sarcopenia and osteoporosis. Ongoing research will certainly find new therapeutic targets for prevention and treatment.

Tissue engineering techniques for the regeneration of large bone defects require sufficient vascularisation of the applied constructs to ensure a sufficient supply of oxygen and nutrients. In our previous work, prevascularised 3D scaffolds have been successfully established by coculture of bone marrow derived stem cells (MSCs) and endothelial progenitor cells (EPCs). We identified stabilising pericytes (PCs) as part of newly formed capillary-like structures. In the present study, we report preliminary data on the interactions between MSCs and EPCs, leading to the differentiation of pericyte-like cells. MSCs and EPCs were seeded in transwell cultures, direct cocultures, and single cultures. Cells were cultured for 10 days in IMDM 10% FCS or IMDM 5% FCS 5% platelet lysate medium. Gene expression of PC markers, CD146, NG2, αSMA, and PDGFR-β, was analysed using RT-PCR at days 0, 3, 7, and 10. The upregulation of CD146, NG2, and αSMA in MSCs in direct coculture with EPCs advocates the MSCs' differentiation towards a pericyte-like phenotype in vitro. These results suggest that pericyte-like cells derive from MSCs and that cell-cell contact with EPCs is an important factor for this differentiation process. These findings emphasise the concept of coculture strategies to promote angiogenesis for cell-based tissue engineered bone grafts.

The gold standard for the treatment of critical-size bone defects is autologous or allogenic bone graft. This has several limitations including donor site morbidity and the restricted supply of graft material. Cell-based tissue engineering strategies represent an alternative approach. Mesenchymal stem cells (MSCs) have been considered as a source of osteoprogenitor cells. More recently, focus has been placed on the use of endothelial progenitor cells (EPCs), since vascularization is a critical step in bone healing. Although many of these approaches have demonstrated effectiveness for bone regeneration, cell-based therapies require time consuming and cost-expensive in vitro cell expansion procedures. Accordingly, research is becoming increasingly focused on the homing and stimulation of native cells. The stromal cell-derived factor-1 (SDF-1) – CXCR4 axis has been shown to be critical for the recruitment of MSCs and EPCs. Vascular endothelial growth factor (VEGF) is a key factor in angiogenesis and has been targeted in many studies. Here, we present an overview of the different approaches for delivering homing factors to the defect site by absorption or incorporation to biomaterials, gene therapy, or via genetically manipulated cells. We further review strategies focusing on the stimulation of endogenous cells to support bone repair. Finally, we discuss the major challenges in the treatment of critical-size bone defects and fracture non-unions.

Background: Platelet-rich plasma (PRP) refers to an enriched platelet suspension in plasma. In addition to the clinical application of PRP in the context of various orthopedic diseases and beyond, PRP and platelet lysate (PL) have been in focus in the field of tissue engineering. In this review, we discuss the application of PRP as a cell culture supplement and as part of tissue engineering strategies, particularly emphasizing current hurdles and ambiguities regarding the efficacy of PRP in these approaches. Summary: As a putative autologous replacement for animal-derived supplements such as fetal calf serum (FCS), PRP has been applied as cell culture supplement for the expansion of stem and progenitor cells for tissue engineering applications and cell therapies. Attributed to the high content of growth factors in platelets, PRP has been shown to promote cell growth, which was mostly superior to standard cultures supplemented with FCS, while the differentiation capacity of progenitor cells seems not to be affected. However, it was also suggested that cultivation of cells with PRP significantly alters the protein expression profile in cells in comparison to FCS, indicating that the influence of PRP on cell behavior should be thoroughly investigated. Moreover, different PRP preparation methods and donor variations have to be considered for the use of PRP under good manufacturing practice conditions. PRP has been used for various tissue engineering applications in the context of bone, cartilage, skin, and soft tissue repair, where most studies were conducted in the field of bone tissue engineering. These approaches take either advantage of the release of chemoattractive, angiogenic, proliferative, and putatively pro-regenerative growth factors from PRP, and/or the hydrogel properties of activated PRP, making it suitable as a cell delivery vehicle. In many of these studies, PRP is combined with biomaterials, cells, and in some cases recombinant growth factors. Although the experimental design often does not allow conclusions on the pro-regenerative effect of PRP itself, most publications report beneficial effects if PRP is added to the tissue-engineered construct. Furthermore, it was demonstrated that the release of growth factors from PRP may be tailored and controlled when PRP is combined with materials able to capture growth factors. Key Messages: Platelet-derived preparations such as PRP and PL represent a promising source of autologous growth factors, which may be applied as cell culture supplement or to promote regeneration in tissue-engineered constructs. Furthermore, activated PRP is a promising candidate as an autologous cell carrier. However, the studies investigating PRP in these contexts often show conflicting results, which most likely can be attributed to the lack of standardized preparation methods, particularly with regard to the platelet content and donor variation of PRP. Ultimately, the use of PRP has to be tailored for the individual application.