Biohybrid Bovine Bone Matrix for Controlled Release of Mesenchymal Stem/Stromal Cell Lyosecretome: A Device for Bone Regeneration

Abstract

SmartBone® (SB) is a biohybrid bone substitute advantageously proposed as a class III medical device for bone regeneration in reconstructive surgeries (oral, maxillofacial, orthopedic, and oncology). In the present study, a new strategy to improve SB osteoinductivity was developed. SB scaffolds were loaded with lyosecretome, a freeze-dried formulation of mesenchymal stem cell (MSC)-secretome, containing proteins and extracellular vesicles (EVs). Lyosecretome-loaded SB scaffolds (SBlyo) were prepared using an absorption method. A burst release of proteins and EVs (38% and 50% after 30 min, respectively) was observed, and then proteins were released more slowly with respect to EVs, most likely because they more strongly adsorbed onto the SB surface. In vitro tests were conducted using adipose tissue-derived stromal vascular fraction (SVF) plated on SB or SBlyo. After 14 days, significant cell proliferation improvement was observed on SBlyo with respect to SB, where cells filled the cavities between the native trabeculae. On SB, on the other hand, the process was still present, but tissue formation was less organized at 60 days. On both scaffolds, cells differentiated into osteoblasts and were able to mineralize after 60 days. Nonetheless, SBlyo showed a higher expression of osteoblast markers and a higher quantity of newly formed trabeculae than SB alone. The quantification analysis of the newly formed mineralized tissue and the immunohistochemical studies demonstrated that SBlyo induces bone formation more effectively. This osteoinductive effect is likely due to the osteogenic factors present in the lyosecretome, such as fibronectin, alpha-2-macroglobulin, apolipoprotein A, and TGF-β.

Effect of Different Bone Grafting Materials and Mesenchymal Stem Cells on Bone Regeneration: A Micro-Computed Tomography and Histomorphometric Study in a Rabbit Calvarial Defect Model

“…Moreover, achieving a balance between the resorption of bone grafting materials during the tissue remodeling process and the maintenance of the bone defect volume for new bone ingrowth will improve osteoinduction [ 6 ]. For example, Smartbone ® , a xenohybrid bone graft, is commercially available as a CE-labeled class III medical device and has shown high levels of bioactivity when loaded with MSCs and lyosecretome [ 5 , 7 ].…”

“…Tissue engineering involves the application of biological and engineering principles for the repair and functional enhancement of human tissues [ 1 ]. In particular, for the reconstruction of craniofacial bone defects, interdisciplinary methods and concepts, including the use of suitable scaffold materials, feasible seed cells, secretome, and bioactive factors, are considered to be vital in the field of bone regeneration [ 2 , 3 , 4 , 5 , 6 , 7 ]. Although the autogenous bone is considered the gold standard for the reconstruction of bone defects, several disadvantages limit its clinical application, including the morbidity of the potential donor site, the requirement of additional surgery, and the low availability of a suitable autologous material [ 8 , 9 ].…”

“…On the other hand, a combination of synthetic and xenograft bone grafting materials (Smartbone ® ) has been proposed and developed. Smartbone ® is a biohybrid bone substitute constituted of a bovine bone-derived matrix and a thin poly(L-lactide- co-ε-caprolactone) (PLCL) film integrated with RGD-containing collagen fragments, which can enhance the cell viability, cell adhesion, and biocompatibility [ 7 ].Therefore, stem cell-based therapy has emerged as an alternative strategy in bone tissue engineering to overcome the limitations of synthetic bone grafting materials and xenografts.…”

“…The efficacy of MSC-based therapy may be affected by the characteristics of bone grafting materials [ 11 , 32 ]. Previous in vivo studies have demonstrated that collagen scaffolds enriched with periodontal ligament-derived MSCs (PDLSCs) or MSC-derived secretome [ 4 ] and a xenohybrid bone graft (Smartbone ® ) combined with adipose-derived stromal vascular fraction or lyoseretome can effectively induce new bone formation, stimulate the recruitment of endogenous bone marrow MSCs, and promote an osteoinductive ability [ 5 , 7 ].…”

The Role of MSCs and Cell Fusion in Tissue Regeneration

“…Limitations include its availability and morbidity, which is why the use of MSCs and other natural or synthetic bone substitutes, as well as the combination of both has been further explored over time. One of those materials is termed SmartBone®, a biohybrid bone substitute [46]. Bari et al used this scaffold and the lyosecretome, consisting of EVs and proteins, of MSCs to show that the lyosecretome improved bone formation [46].…”