Identification of mesenchymal stem cells and osteogenic factors in bone marrow aspirate and peripheral blood for spinal fusion by flow cytometry and proteomic analysis

Abstract

BackgroundAn in vivo animal study and a prospective clinical study have indicated that bone marrow aspirate (BMA) augments spinal arthrodesis. However, there is no quantified data to explain why fusion rate can be augmented by BMA in lumbar posterolateral fusion.MethodsTo analyze the proportion of mesenchymal stem cells (MSCs) and osteogenic factors in human BMA and peripheral blood (PB) of the same patient. Autologous BMA and PB from the patients were analyzed by flow cytometry (FACS) using cell markers for MSCs. The osteogenic potential of MSCs was determined by alkaline phosphatase (ALP) activity and calcium level quantification. Proteomics were used for the qualitative and quantitative mapping of the whole proteome from BMA and PB plasma. The mass-to-charge ratio was calculated by time-of-flight mass spectrometry (TOF-MS). The overexpression of protein was confirmed using Western blot analysis.ResultsThe proportion of MSCs (CD34−/CD29+/CD105+) was higher in the BMA than that in the PB. Colony-forming cell (CFC) assays suggested that fewer colonies were formed in PB cultures than in BMA culture. There was no significant difference in the osteogenic potential of the MSCs between the PB and BMA. Proteomic mass spectrometry assays suggested that the levels of catalase (osteoclast inhibitor) and glutathione peroxidase 3 (osteogenic biomarker) were higher in the BMA than those in the PB, and this was confirmed by Western blot analysis.ConclusionsThe proportions of MSCs and osteogenic factors were higher in the BMA than in the PB. This may explain why fusion rate can be augmented by BMA in lumbar posterolateral fusion.

Heparin Anticoagulant for Human Bone Marrow Does Not Influence In Vitro Performance of Human Mesenchymal Stromal Cells

“…Thirdly, the applied dosages vary largely. Some studies indicate that mere rinsing of syringes with heparin solution was applied [3] but others used 100 IU/mL bone marrow [4]. Two types of heparins are widely used, high molecular weight unfractionated heparin and low molecular weight fractionated heparin.…”

Bone marrow coagulated and low‐level laser therapy accelerate bone healing by enhancing angiogenesis, cell proliferation, osteoblast differentiation, and mineralization

“…However, the same BMA harvest protocol was performed by an experienced professional, which is very important considering that the quality of BMA is extremely technique‐dependent 9 . Flow cytometer results did show that BMA samples express cell surface markers related to MSC, hematopoietic, and endothelial cells as observed in previous studies 33‐35 . So, it can be inferred that the improvement on bone healing was mainly due to the presence of MSC, hematopoietic stem cells, and endothelial cells at the wound site.…”

Understanding the Future Prospects of Synergizing Minimally Invasive Transforaminal Lumbar Interbody Fusion Surgery with Ceramics and Regenerative Cellular Therapies

“…In recent decades, the differentiation potential of stem cells has extensively been explored to develop regenerative therapies for various disorders. It has been reported that as a bone graft, the bone marrow aspirate (BMA) rich in mesenchymal stem cells (MSCs) and osteogenic factor could efficaciously stimulate spinal bone fusion [ 176 , 177 ]. The osteogenic potential of bone marrow stem cells (BMSCs) also plays a key supportive role in the therapeutic effect of BMA in bone fusion.…”