Hemovac blood after total knee arthroplasty as a source of stem cells

Abstract

Background With increasing life expectancy, stem cell therapy is receiving increasing attention. However, its application is restricted by ethical concerns. Hence a need exists for design of safe procedures for stem cell procurement. Here, we investigated whether hemovac blood (HVB) is an appropriate stem cell source. Methods HVB concentrates (HVBCs) from 20 total knee arthroplasty (TKA) patients and bone marrow aspirate (BMA) concentrates (BMACs) from 15 patients who underwent knee cartilage repair were comparatively evaluated. A bone marrow aspiration needle was inserted into the anterior superior iliac spine. Aspiration was performed using a 50-mL syringe, including 4 mL of anticoagulant, followed by centrifugation to obtain BMACs. To obtain HVBCs, blood was aspirated from the hemovac immediately after TKA surgery. Different cell types were enumerated. Isolation of BMA and HVB mononuclear cells was performed using density gradient centrifugation. Non-hematopoietic fibroblast colonies were quantified by colony forming unit-fibroblast assay surface marker analysis of HVB, HVBC, BMA, and BMAC was performed via flow cytometry. Mesenchymal stem cells (MSCs) isolated from HVBCs and BMACs were examined for osteogenic, adipogenic, and chondrogenic differentiation potential. Gene expression analysis was performed by quantitative real-time polymerase chain reaction (qRT-PCR). Results The number of cells from HVB and HVBC was significantly lower than from BMA and BMAC; however, the number of colonies in HVBC and BMAC did not differ significantly (P>0.05). Isolated cells from both sources had a fibroblast-like appearance, adhered to culture flasks, and formed colonies. Under different culture conditions, MSC-specific surface markers (CD29, CD44, CD90, CD105), osteogenic markers [RUNX2, osteopontin, osteocalcin, and alkaline phosphatase (ALP)] and adipogenic markers (PPARγ and C/EBPα) were expressed. Moreover, SOX9, type II collagen, and aggrecan were significantly upregulated upon chondrogenic differentiation. Conclusions HVB from TKA patients is a useful source of stem cells for research.