Coculture of Peripheral Blood-Derived Mesenchymal Stem Cells and Endothelial Progenitor Cells on Strontium-Doped Calcium Polyphosphate Scaffolds to Generate Vascularized Engineered Bone

Abstract

Vascularization of engineered bone tissue is critical for ensuring its survival after implantation and it is the primary factor limiting its clinical use. A promising approach is to prevascularize bone grafts in vitro using endothelial progenitor cells (EPC) derived from peripheral blood. Typically, EPC are added together with mesenchymal stem cells (MSC) that differentiate into osteoblasts. One problem with this approach is how to promote traditional tissue engineering bone survival with a minimally invasive method. In this study, we examined the effectiveness of administering to stimulate the release of peripheral blood stem cells and their co-culturing system for generating prevascularized engineered bone. Cells were isolated by Ficoll density gradient centrifugation and identified as EPC and MSC based on morphology, surface markers, and functional analysis. EPC and MSC were cocultured in several different ratios, and cell morphology and tube formation were assessed by microscopy. Expression of osteogenesis and vascularization markers was quantified by enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction, and histochemical and immunofluorescence staining. Increasing the proportion of EPC in the coculture system led to greater tube formation and greater expression of the endothelial cell marker CD31. An EPC:MSC ratio of 75:25 gave the highest expression of osteogenesis and angiogenesis markers. Cocultures adhered to a three-dimensional scaffold of strontium-doped calcium polyphosphate and proliferated well. Our findings show that coculturing peripheral blood-derived EPC and MSC may prove useful for generating prevascularized bone tissue for clinical use.

Cocultivation of Mesenchymal Stem Cells and Endothelial Progenitor Cells Reveals Antiapoptotic and Proangiogenic Effects

“…Knowing that endothelial cells and EPCs can induce the osteogenic differentiation of MSCs [Thébaud et al, 2012;Fu et al, 2015], we measured the early osteogenic differentiation marker ALP as well as the concentration of the osteogenic growth factor BMP-2. In the pertinent literature, ALP is described as an early osteogenic differentiation marker.…”

Overcoming physical constraints in bone engineering: ‘the importance of being vascularized’

“…The key contribution of cell milieu is supported by a great deal of research, part of which related to the sophisticated mimicking of structural and functional details of the extracellular matrix with artificial of semi-synthetic materials. (Cao et al, 2014;Fu et al, 2014;Shim, Ankeny, Kim, Nerem, & Khang, 2014;Tanase et al, 2013;Yeatts et al, 2014;Q. Zhao et al, 2010).…”

“…The feasibility of co-culture or co-implantation strategies has been so far tested in several studies, focusing on the quality of the engineered bone substitute (Dariima, Jin, Lee, Wall, & Kim, 2013;Fu et al, 2014;Kokemüller et al, 2014;Leszczynska et al, 2013;R P Pirraco et al, 2013;Rogério P Pirraco et al, 2014). Interestingly, mineralization can be achieved without osteogenic media or additional growth factors through co-cultures of normal human primary osteoblats and Human umbilical vein ECs (HUVECs) mainly by recurring to the manipulation of the coculture ratio (Shah, Wenke, & Agrawal, 2014).…”

Cell Sheets of Co-cultured Endothelial Progenitor Cells and Mesenchymal Stromal Cells Promote Osseointegration in Irradiated Rat Bone

“…Studies demonstrated that co-cultured EPCs and BMSCs could enhance bone regeneration 14, 24, 25 . However, no consensus on the condition for co-culture has been achieved 26 . In order to determine the best ratio of co-culture for bone regeneration, we co-cultured EPCs and BMSCs at ratios of 1:0, 10:1, 5:1, 1:1, 1:5, 1:10 and 0:1.…”