Oxygen delivery augmented bone formation from transplanted bone marrow

Abstract

Event Abstract Back to Event Oxygen delivery augmented bone formation from transplanted bone marrow Huaifa Zhang1, Daisuke Sato1, Uwe Gbureck2, Benjamin Dalisson1, Simon Tran1 and Jake Barralet1, 3 1 McGill University, Faculty of Dentistry, Canada 2 Julius Maximilians University of Wuerzburg, Department of Functional Materials in Medicine and Dentistry, Germany 3 McGill University, Division of Orthopaedics, Department of Surgery, Faculty of Medicine, Canada Introduction: Bone marrow contains cells that are able to differentiate into bone. Ectopically transplanted bone marrow has also been found to generate bone tissue in vivo[1]. However, hypoxia occurs after tissue transplantation due to the limited blood supply into the transplant. As a result, much of the transplanted marrow tissue dies, thereby reducing the bone regeneration ability of marrow tissue. So far, no effective ways have been reported to reduce cell death in marrow tissue post transplantation. Calcium phosphate scaffolds have been widely used for bone regeneration because of their good osteoconductivity. It has been found that bone formation happens when cultured bone marrow-derived cells within calcium phosphate scaffolds are transplanted[2]. We have developed a self-oxygenating scaffold consisting of metal peroxides and alginate hydrogel. In this work, we encapsulated rat bone marrow into the hydrogel and cultured rat bone marrow under anoxia to determine the potential for survival post implantation. Afterwards, we added the encapsulated rat bone marrow into a 3D printed closed ended calcium phosphate cylindrical scaffold. In the end, we transplanted the scaffold containing marrow tissue subcutaneously in a rat model. Experimental: The bone marrow from rat tibia and femurs was harvested, mixed with sodium alginate and further cross-linked with CaCl2. The encapsulated marrow tissue with and without oxygen release material was cultured under anoxia (95% N2, 5% CO2, 37°C). Cell viability in cultured bone marrow was examined with live/dead and LDH assays. Afterwards, the encapsulated bone marrow was incorporated into the calcium phosphate scaffold and transplanted subcutaneously into rats. Six implants were implanted in each rat (dorsal side), three implants each side. After two and four weeks, the implants were retrieved and fixed. Bone formation in the implanted scaffolds was studied with Micro-CT. Then the scaffolds were embedded in paraffin and resin, respectively, for histological characterization. Result and Discussion: Figure 1 shows the live and dead cells in the marrow. It was apparent that the oxygen release scaffold drastically improved cell viability under anoxia. In the control experiment, most of the cells died after seven days under anoxia, whereas the tissue cultured with oxygen release scaffold exhibited very high cell viability (around 90%). In vivo experimental results show a larger amount of minerals in the presence of oxygen release material compared to the control group (Figure 2). The Micro-CT images clearly show that there were more minerals in the core area of the implants in the experimental group compared to the control group. The newly formed minerals was about 2.2 v/v% and 1 v/v% in experimental group and control group, respectively, after two weeks. Figure 1: Fluorescent images of live/dead assay stained marrow tissue cultured in anoxia with (Left) and without (Right) oxygen release material after seven days. Live cells were stained into green and dead cells into red. The nuclei of all cells were stained into blue. Figure 2 : Micro-CT images of retrieved marrow implants with (Left) and without (Right) oxygen release material after two weeks. Conclusion: To the best of our knowledge, this is the first attempt ever made to sustain bone marrow tissue in vitro using oxygen delivery material. We successfully developed a self-oxygenating scaffold which is capable of improved cell viability in bone marrow tissue under anoxia. Furthermore, the oxygen release scaffold augmented mineral formation after subcutaneous implantation. This is of great clinical interest and offers new prospects in improving clinical applications of bone marrow transplantation. The Natural Sciences and Engineering Research Council of Canada; China Scholarship Council