Harnessing extracellular vesicles to direct endochondral repair of large bone defects

Abstract

Large bone defects remain a tremendous clinical challenge. There is growing evidence in support of treatment strategies that direct defect repair through an endochondral route, involving a cartilage intermediate. While culture-expanded stem/progenitor cells are being evaluated for this purpose, these cells would compete with endogenous repair cells for limited oxygen and nutrients within ischaemic defects. Alternatively, it may be possible to employ extracellular vesicles (EVs) secreted by culture-expanded cells for overcoming key bottlenecks to endochondral repair, such as defect vascularization, chondrogenesis, and osseous remodelling. While mesenchymal stromal/stem cells are a promising source of therapeutic EVs, other donor cells should also be considered. The efficacy of an EV-based therapeutic will likely depend on the design of companion scaffolds for controlled delivery to specific target cells. Ultimately, the knowledge gained from studies of EVs could one day inform the long-term development of synthetic, engineered nanovesicles. In the meantime, EVs harnessed from in vitro cell culture have near-term promise for use in bone regenerative medicine. This narrative review presents a rationale for using EVs to improve the repair of large bone defects, highlights promising cell sources and likely therapeutic targets for directing repair through an endochondral pathway, and discusses current barriers to clinical translation. Cite this article: E. Ferreira, R. M. Porter. Harnessing extracellular vesicles to direct endochondral repair of large bone defects. Bone Joint Res 2018;7:263-273. DOI: 10.1302/2046-3758.74.BJR-2018-0006.

Keywords: Critical-sized bone defects; Endochondral ossification; Extracellular vesicles; Mesenchymal stromal cells.

Fig. 1

Diagram showing intercellular communication by extracellular vesicles (EVs). Two principle EV fractions are understood to play roles in intercellular communication: exosomes, which are released from multivesicular bodies after fusing with the parent cell plasma membrane; and larger microvesicles, which bud directly from the parent cell membrane. Each fraction contains unique profiles of intravesicular RNAs and protein as well as membrane-bound receptors and lipids. These fractions stimulate responses within recipient cells by direct activation of recipient cell surface receptors, by transfer of vesicle contents to the recipient cell cytosol after fusion with the plasma membrane, and by intracellular trafficking of vesicle contents following endocytosis.

Fig. 2

Directing endochondral repair of large bone defects. One paradigm for bone regenerative medicine is modelled on the processes of long bone development and successful (fracture) repair. Instead of designing scaffold/biological constructs for the direct stimulation of osteogenesis, constructs can be engineered to undergo an initial chondrogenesis phase, which serves as an efficient template for ordered osteogenic remodelling by successive waves of repair cells. It is noteworthy that cartilage, an avascular tissue, is more resilient to the vascular deficiency within larger bone defects.

Fig. 3

Diagram showing the potential advantage of extracellular vesicles (EVs) within a large bone defect microenvironment. As opposed to simple fractures, bone defects beyond a critical size are characterized by severe nutrient deficiency and near-anoxia within their core. While exogenous cells implanted into these defects may secrete pro-regenerative factors, they also compete with endogenous repair cells migrating into the defect for scarce oxygen and nutrients. In contrast, the same pro-regenerative signals packaged within EVs would not necessarily tax the defect for nutrients and oxygen, potentially permitting enhanced repair by endogenous cells.

Fig. 4

Diagram showing therapeutic targets for endochondral repair. Bottlenecks to endochondral bone repair include progressive vascularization, chondroprogenitor recruitment, neocartilage formation, and osseous remodelling. It may be possible to deliver extracellular vesicles (EVs) harvested in vitro from promising parent cell cultures that stimulate endogenous cells to overcome one or more of these bottlenecks. An ideal scaffold for this approach would not only be chondro-conductive but would also control the release of therapeutic EVs, in order to match the migration timeframe of target repair cells (endothelial, chondroprogenitor) into the defect.

Fig. 5

Diagram showing the genetic engineering of extracellular vesicles (EVs) for targeting endogenous repair cells. The efficacy of exogenous EVs may be improved by introducing targeting ligands onto their surface that recognize cell surface receptors specific for key repair cells. One way in which to introduce these ligands would be to genetically engineer parent cells to express them within the extracellular domain of a membrane-anchored fusion protein. The fusion protein would then be expressed on the surface of EVs secreted by the parent cells. In addition to potentially improving the efficiency of target cell uptake of EV contents, this strategy could also limit nonspecific or adverse effects in off-target cells.