Functionalized 3D-Printed ST2/Gelatin Methacryloyl/Polcaprolactone Scaffolds for Enhancing Bone Regeneration with Vascularization

Abstract

Growth factors were often used to improve the bioactivity of biomaterials in order to fabricate biofunctionalized bone grafts for bone defect repair. However, supraphysiological concentrations of growth factors for improving bioactivity could lead to serious side effects, such as ectopic bone formation, radiculitis, swelling of soft tissue in the neck, etc. Therefore, safely and effectively applying growth factors in bone repair biomaterials comes to be an urgent problem that needs to be addressed. In this study, an appropriate concentration (50 ng/mL) of Wnt3a was used to pretreat the 3D-bioprinting gelatin methacryloyl(GelMA)/polycaprolactone(PCL) scaffold loaded with bone marrow stromal cell line ST2 for 24 h. This pretreatment promoted the cell proliferation, osteogenic differentiation, and mineralization of ST2 in the scaffold in vitro, and enhanced angiogenesis and osteogenesis after being implanted in critical-sized mouse calvarial defects. On the contrary, the inhibition of Wnt/β-catenin signaling in ST2 cells reduced the bone repair effect of this scaffold. These results suggested that ST2/GelMA/PCL scaffolds pretreated with an appropriate concentration of Wnt3a in culture medium could effectively enhance the osteogenic and angiogenic activity of bone repair biomaterials both in vitro and in vivo. Moreover, it would avoid the side effects caused by the supraphysiological concentrations of growth factors. This functionalized scaffold with osteogenic and angiogenic activity might be used as an outstanding bone substitute for bone regeneration and repair.

CHIR99021-Treated Osteocytes with Wnt Activation in 3D-Printed Module Form an Osteogenic Microenvironment for Enhanced Osteogenesis and Vasculogenesis

“…In our laboratory, we found that Wnt-activated osteocytes may affect bone formation by influencing surrounding skeletal cells through paracrine signaling, and specific data showed that Wnt-activated osteocytes promote bone formation, bone resorption [ 15 ], and stem cell proliferation. These findings have translational applications [ 21 , 31 , 32 ] and will be beneficial for BTE. We found that C91 effectively prevented the degradation of β-catenin in the cytoplasm of MLO-Y4 osteocytes and mediated β-catenin entry into the nucleus, thereby significantly increasing the expression of Wnt target genes.…”

“…In the early stages of our research, we found that the bone cell acellular matrix that was activated by Wnt signaling via gene manipulation accelerated the repair of critical-sized parietal bone defects by promoting osteoblast formation, angiogenesis, and neurogenesis [ 21 ]. Moreover, GelMA/PCL scaffolds containing ST2 cells pretreated with Wnt3a can effectively enhance the osteogenic and angiogenic activities of bone repair biomaterials in vitro and in vivo [ 32 ]. Our team has proven that this functional module has osteogenic and vascular functions, solves the problem of biological activity, and can be used as a good alternative material for bone regeneration and bone repair.…”

“…Alkaline phosphatase staining (AP staining) was performed as previously described [ 31 , 32 ]. After 3 days of cell culture and aspirating the medium, the cells were washed with PBS (Sorlabio, Beijing, China) two times to remove any interference with proteins contained in the serum.…”

“…The AP biochemical activity assay was executed as previously reported [ 31 , 32 ]. After aspirating the medium and washing the cells with PBS, 300 µL 10 mM Tris/HCl (pH 7.4) was added to each well.…”

Three-Dimensional Cell Cultures: The Bridge between In Vitro and In Vivo Models

“…On the other side, several in vivo studies have reported that the use of composite biomaterials, such as HA/alginate or HA/collagen, improved, by approximately fourfold, the regenerative potential of endogenous MSC cells by increasing bone formation [357]. Similarly, other studies have reported that 3D systems functionalized with growth factors [358,359], angiogenic factors [360], miRNA [361], exosomes [362], antibiotics [363], and other compounds are considered to be an added value in promoting bone regeneration [358]. Several challenges remain related to the application of cell therapy or the use of tissue-engineered or functionalized scaffolds to promote bone regeneration in vivo.…”