Optimal concentration of mesenchymal stem cells for fracture healing in a rat model with long bone fracture

Myung-Seo Kim; Hyun-Ju Chung; Kang-Il Kim

doi:10.4252/wjsc.v14.i12.839

Optimal concentration of mesenchymal stem cells for fracture healing in a rat model with long bone fracture

World J Stem Cells. 2022 Dec 26;14(12):839-850. doi: 10.4252/wjsc.v14.i12.839.

Authors

Myung-Seo Kim¹, Hyun-Ju Chung², Kang-Il Kim³

Affiliations

¹ Department of Orthopaedic Surgery, School of Medicine, Kyung Hee University and Kyung Hee University Hospital at Gangdong, Seoul 05278, South Korea.
² Department of Core Research Laboratory, Clinical Research Institute, Kyung Hee University Hospital at Gangdong, Seoul 05278, South Korea.
³ Department of Orthopaedic Surgery, School of Medicine, Kyung Hee University and Kyung Hee University Hospital at Gangdong, Seoul 05278, South Korea. khuknee@gmail.com.

Abstract

Background: There is still no consensus on which concentration of mesenchymal stem cells (MSCs) to use for promoting fracture healing in a rat model of long bone fracture.

Aim: To assess the optimal concentration of MSCs for promoting fracture healing in a rat model.

Methods: Wistar rats were divided into four groups according to MSC concentrations: Normal saline (C), 2.5 × 10⁶ (L), 5.0 × 10⁶ (M), and 10.0 × 10⁶ (H) groups. The MSCs were injected directly into the fracture site. The rats were sacrificed at 2 and 6 wk post-fracture. New bone formation [bone volume (BV) and percentage BV (PBV)] was evaluated using micro-computed tomography (CT). Histological analysis was performed to evaluate fracture healing score. The protein expression of factors related to MSC migration [stromal cell-derived factor 1 (SDF-1), transforming growth factor-beta 1 (TGF-β1)] and angiogenesis [vascular endothelial growth factor (VEGF)] was evaluated using western blot analysis. The expression of cytokines associated with osteogenesis [bone morphogenetic protein-2 (BMP-2), TGF-β1 and VEGF] was evaluated using real-time polymerase chain reaction.

Results: Micro-CT showed that BV and PBV was significantly increased in groups M and H compared to that in group C at 6 wk post-fracture (P = 0.040, P = 0.009; P = 0.004, P = 0.001, respectively). Significantly more cartilaginous tissue and immature bone were formed in groups M and H than in group C at 2 and 6 wk post-fracture (P = 0.018, P = 0.010; P = 0.032, P = 0.050, respectively). At 2 wk post-fracture, SDF-1, TGF-β1 and VEGF expression were significantly higher in groups M and H than in group L (P = 0.031, P = 0.014; P < 0.001, P < 0.001; P = 0.025, P < 0.001, respectively). BMP-2 and VEGF expression were significantly higher in groups M and H than in group C at 6 wk post-fracture (P = 0.037, P = 0.038; P = 0.021, P = 0.010). Compared to group L, TGF-β1 expression was significantly higher in groups H (P = 0.016). There were no significant differences in expression levels of chemokines related to MSC migration, angiogenesis and cytokines associated with osteogenesis between M and H groups at 2 and 6 wk post-fracture.

Conclusion: The administration of at least 5.0 × 10⁶ MSCs was optimal to promote fracture healing in a rat model of long bone fractures.

Keywords: Direct injection; Femoral shaft fracture; Fracture healing; Mesenchymal stem cells; Optimal concentration; Rat model.