Repair of complex fractures with bone loss requires a potent, space-filling intervention to promote regeneration of bone. We present a biomaterials-based strategy combining mesenchymal stromal cells (MSC) with a chitosan-collagen matrix to form modular microtissues designed for delivery through a needle to conformally fill cavital defects. Implantation of microtissues into a calvarial defect in the mouse showed that osteogenically pre-differentiated MSC resulted in complete bridging of the cavity, while undifferentiated MSC produced mineralized tissue only in apposition to native bone. Decreasing the implant volume reduced bone regeneration, while increasing the MSC concentration also attenuated bone formation, suggesting that the cell-matrix ratio is important in achieving a robust response. Conformal filling of the defect with microtissues in a carrier gel resulted in complete healing. Taken together, these results show that modular microtissues can be used to augment the differentiated function of MSC and provide an extracellular environment that potentiates bone repair.
Inadequate vascularization of engineered tissue constructs is a main challenge in developing a clinically impactful therapy for large, complex, and recalcitrant bone defects. It is well established that bone and blood vessels form concomitantly during development, as well as during repair after injury. Endothelial cells (ECs) and mesenchymal stromal cells (MSCs) are known to be key players in orthopedic tissue regeneration and vascularization, and these cell types have been used widely in tissue engineering strategies to create vascularized bone. Coculture studies have demonstrated that there is crosstalk between ECs and MSCs that can lead to synergistic effects on tissue regeneration. At the same time, the complexity in fabricating, culturing, and characterizing engineered tissue constructs containing multiple cell types presents a challenge in creating multifunctional tissues. In particular, the timing, spatial distribution, and cell phenotypes that are most conducive to promoting concurrent bone and vessel formation are not well understood. This review describes the processes of bone and vascular development, and how these have been harnessed in tissue engineering strategies to create vascularized bone. There is an emphasis on interactions between ECs and MSCs, and the culture systems that can be used to understand and control these interactions within a single engineered construct. Developmental engineering strategies to mimic endochondral ossification are discussed as a means of generating vascularized orthopedic tissues. The field of tissue engineering has made impressive progress in creating tissue replacements. However, the development of larger, more complex, and multifunctional engineered orthopedic tissues will require a better understanding of how osteogenesis and vasculogenesis are coupled in tissue regeneration.
Bioengineered bone designed to heal large defects requires concomitant development of osseous and vascular tissue to ensure engraftment and survival. Adult human mesenchymal stromal cells (MSC) are promising in this application because they have demonstrated both osteogenic and vasculogenic potential. This study employed a modular approach in which cells were encapsulated in biomaterial carriers (microtissues) designed to support tissue‐specific function. Osteogenic microtissues consisting of MSC embedded in a collagen‐chitosan matrix; vasculogenic (VAS) microtissues consisted of endothelial cells and MSC in a fibrin matrix. Microtissues were precultured under differentiation conditions to induce appropriate MSC lineage commitment, and were then combined in a surrounding fibrin hydrogel to create a multimodular construct. Results demonstrated the ability of microtissues to support lineage commitment, and that preculture primes the microtissues for the desired function. Combination of osteogenic and vasculogenic microtissues into multimodular constructs demonstrated that osteogenic priming resulted in sustained osteogenic activity even when cultured in vasculogenic medium, and that vasculogenic priming induced a pericyte‐like phenotype that resulted in development of a primitive vessel network in the constructs. The modular approach allows microtissues to be separately precultured to harness the dual differentiation potential of MSC to support both bone and blood vessel formation in a unified construct.
<150 Words):Repair of complex fractures with bone loss requires a potent, space-filling intervention to promote regeneration of bone. We present a minimally-invasive strategy combining mesenchymal stromal cells (MSC) with a chitosan-collagen matrix to form modular microtissues designed for delivery through a needle to conformally fill cavital defects. Implantation of microtissues into a calvarial defect in the mouse showed that osteogenically pre-differentiated MSC resulted in complete bridging of the cavity, while undifferentiated MSC produced mineralized tissue only in apposition to native bone. Decreasing the implant volume reduced bone regeneration, while increasing the MSC concentration also attenuated bone formation, suggesting that the cell-matrix ratio is important in achieving a robust response. Conformal filling of the defect with microtissues in a carrier gel resulted in complete healing. Taken together, these results show that modular microtissues can be used to augment the differentiated function of MSC and provide an extracellular environment that potentiates bone repair. Introduction:Bone has a remarkable capacity to regenerate through carefully orchestrated, cell-mediated repair processes [1]. However, healing in large and complex fractures is often impaired, leading to incomplete or functionally inferior bone regeneration. In some wounds, loss of the native vasculature and infection of the wound bed can further impair bone regeneration, resulting in a variety of pathologies, including delayed-, mal-, and non-unions. In such cases, therapeutic intervention to stimulate and accelerate the healing response is required. Bone grafting is a standard approach to this problem but needs invasive surgery to harvest and deliver the graft.Autologous grafts and flaps are limited in supply and can cause donor-site morbidity, including infection, hematoma, and pain [2]. Moreover, they are not suitable in 10-30% of cases due to difficulty in conforming the graft to the shape of the defect [3]. Allogeneic decellularized grafts and synthetic ceramic substitutes can also be used, but are biologically inferior compared to viable bone grafts due to the lack of cellular components. Although processes such as irradiation and lyophilization can reduce the risk of disease transmission from an allogeneic graft, they eliminate cellular components resulting in reduced osteoinductivity [4] and revascularization, resulting in higher bone resorption [5].The ideal bone substitute would exhibit osteoconductive, osteoinductive, and osteogenic properties and would promote concomitant neovascularization of larger defects [4]. Materialsbased approaches have been developed to promote osteoconductivity [1], and the immobilization and release of growth factors can be used as an osteoinductive cue. However, only cells can produce bone, and osteogenesis, therefore, requires either recruitment of endogenous cells or delivery of exogenous cells capable of forming bone. In large and ischemic defects, endogenous cell recruitment is impaired,...
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