Prevascularization of biofunctional calcium phosphate cement for dental and craniofacial repairs

Abstract

Objectives Calcium phosphate cement (CPC) is promising for dental and craniofacial repairs. Vascularization in bone tissue engineering constructs is currently a major challenge. The objectives of this study were to investigate the prevascularization of macroporous CPC via coculturing human umbilical vein endothelial cells (HUVEC) and human osteoblasts (HOB), and determine the effect of RGD in CPC on microcapillary formation for the first time. Methods Macroporous CPC scaffold was prepared using CPC powder, chitosan liquid and gas-foaming porogen. Chitosan was grafted with Arg-Gly-Asp (RGD) to biofunctionalize the CPC. HUVEC and HOB were cocultured on macroporous CPC-RGD and CPC control without RGD for up to 42 d. The osteogenic and angiogenic differentiation, bone matrix mineral synthesis, and formation of microcapillary-like structures were measured. Results RGD-grafting in CPC increased the genes expressions of osteogenic and angiogenic differentiation markers than those of CPC control without RGD. Cell-synthesized bone mineral content also increased on CPC-RGD, compared to CPC control (p < 0.05). Immunostaining with endothelial marker showed that the amount of microcapillary-like structures on CPC scaffolds increased with time. At 42 d, the cumulative vessel length for CPC-RGD scaffold was 1.69-fold that of CPC control. SEM examination confirmed the morphology of self-assembled microcapillary-like structures on CPC scaffolds. Significance HUVEC+HOB coculture on macroporous CPC scaffold successfully achieved prevascularization. RGD incorporation in CPC enhanced osteogenic differentiation, bone mineral synthesis, and microcapillary-like structure formation. The novel prevascularized CPC-RGD constructs are promising for dental, craniofacial and orthopedic applications.

Angiogenic and osteogenic regeneration in rats via calcium phosphate scaffold and endothelial cell co-culture with human bone marrow mesenchymal stem cells (MSCs), human umbilical cord MSCs, human induced pluripotent stem cell-derived MSCs and human embry

“…CPC has good biocompatibility, osteoconductivity and bone replacement capability. The incorporation of angiogenic growth factors in CPC and the creation of microvascular networks in CPC prior to implantation were previously studied to address the issue of limited angiogenesis and thus insufficient bone formation (Wernike et al , ; Chen et al , ). However, a literature search revealed no report on the investigation of co‐culture of ECs with different types of MSC on bone regeneration efficacy.…”

“…The lengths of microcapillaries weres measured with Image‐Pro Plus software (Media Cybernetics, Silver Springs, MD, USA) to obtain the cumulative length of microcapillaries for each image. The cumulative length of microcapillaries of the image was divided by the area of that image, to yield the cumulative microcapillary length per scaffold surface area (Chen et al , ).…”

“…Gene expressions were measured by TaqMan gene expression kits, including human alkaline phosphatase (ALP, Hs00758162_m1), osteocalcin (OC, Hs00609452_g1), collagen type I (COL I, Hs00164004), vascular endothelial growth factor A (VEGF) (HS00900055_ml), vascular endothelial cadherin (VE‐cadherin) (Hs00170986_m1), von‐Willebrand factor (vWF) (Hs00169795_m1) and glyceraldehyde 3‐phosphate dehydrogenase (GAPDH, Hs99999905). The 2 ‐ΔΔCt method was used to evaluate the relative expression for each target gene (Chen et al , ). The C t of target genes was normalized by the C t of the housekeeping gene GAPDH to obtain Δ C t .…”

A self-setting iPSMSC-alginate-calcium phosphate paste for bone tissue engineering

“…CPC powder was prepared following a previous study [33–34]. Briefly, TTCP [Ca 4 (PO 4 ) 2 O] was synthesized using DCPA (CaHPO 4 ) and calcium carbonate (both from J.T.…”

“…The commercial DCPA powder was ground for 24 h in the ball mill in 95% ethanol and sieved to obtain a median particle size of approximately 1 μm. Then the TTCP and DCPA powders at 1:3 molar ratio were thoroughly mixed in a micromill (Bel-Alert Products, Pequannock, NJ) to form the CPC powder [33–34]. The microbeads were incorporated into CPC at 50% by volume.…”