Abstract
The potential use of bone progenitors, multipotential stromal cells (MSCs) helping spine fusion is increasing, but convenient MSC sources and effective processing methods are critical factors yet to be optimised. The aim of this study was to test the effect of bone marrow processing on the MSC abundance and to compare the differentiation capabilities of vertebral body-bone marrow (VB-BM) MSCs versus iliac crest-bone marrow (IC-BM) MSCs. We assessed the effect of the red blood cell lysis (ammonium chloride, AC) and density-gradient centrifugation (Lymphoprep™, LMP), on the extracted VB-BM and IC-BM MSC numbers. The MSC abundance (indicated by colony counts and CD45lowCD271high cell numbers), phenotype, proliferation and tri-lineage differentiation of VB-BM MSCs were compared with donor-matched IC-BM MSCs. Importantly, the MSC attachment and osteogenesis were examined when VB-BM and IC-BM samples were loaded on a beta-tricalcium phosphate scaffold. In contrast to LMP, using AC yielded more colonies from IC-BM and VB-BM aspirates (p = 0.0019 & p = 0.0201 respectively). For IC-BM and VB-BM, the colony counts and CD45lowCD271high cell numbers were comparable (p = 0.5186, p = 0.2640 respectively). Furthermore, cultured VB-BM MSCs exhibited the same phenotype, proliferative and adipogenic potential, but a higher osteogenic and chondrogenic capabilities than IC-BM MSCs (p = 0.0010 and p = 0.0005 for calcium and glycosaminoglycan (GAG) levels, respectively). The gene expression data confirmed higher chondrogenesis for VB-BM MSCs than IC-BM MSCs, but osteogenic gene expression levels were comparable. When loaded on Vitoss™, both MSCs showed a similar degree of attachment and survival, but a better osteogenic ability was detected for VB-BM MSCs as measured by alkaline phosphatase activity (p = 0.0386). Collectively, the BM processing using AC had more MSC yield than using LMP. VB-BM MSCs have a comparable phenotype and proliferative capacity, but higher chondrogenesis and osteogenesis with or without using scaffold than donor-matched IC-BM MSCs. Given better accessibility, VB-BM could be an ideal MSC source for spinal bone fusion.
Highlights
Bone progenitor cells, multipotential stromal cells (MSCs) are increasingly used for the reparative bone therapy
The samples of vertebral body-bone marrow (VB-Bone marrow (BM)) were aspirated from the vertebral body as illustrated in S1 Fig. The total numbers of yielded cells of iliac crest-bone marrow (IC-BM) and VB-BM aspirates were counted to assess the effect of processing method (Fig 1A, left panel)
Our data showed that the red blood cell lysis is a convenient method for processing VB-BM samples to extract higher yield of MSCs compared to the gradient centrifugation
Summary
Multipotential stromal cells (MSCs) are increasingly used for the reparative bone therapy. Bone marrow (BM) is the best-studied source for MSCs, being used clinically without or after processing to extract pure culture-expanded MSCs [1]. Bone fusion is a conventional method of treating these conditions, it is not devoid of failures [3]. Bone autograft and osteoconductive scaffolds are commonly used. The scaffolds can be enriched with BM, usually from the iliac crest (IC), or cultured MSCs to further enhance bone fusion [4,5,6]. A swift biological fusion ensures better preservation of the initial surgical correction and fewer complications, there remains a considerable rate of pseudoarthrosis with subsequent pain and metalwork failure despite the advances in surgical techniques [7,8,9]
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