[課題7] Controlling Redox State at Transplantation Site Enhances Bone Regeneration
[Abstract]
[Purpose]
In cell-based bone augmentation, dysfunction and apoptosis of transplanted cells can occur owing to the oxidative stress caused by the overproduction of reactive oxygen species1. Therefore, this study aimed to investigate the effect of controlling oxidative stress on bone regeneration using a free radical scavenger – edaravone (EDA)
[Methods]
Bone marrow-derived cells were collected from 4-week-old rats. The effects of EDA on cell viability and osteogenic differentiation with or without hydroperioxide (H2O2) were evaluated. RNA sequencing was performed to detect differentially expressed genes. Two critical-sized calvaria defects were created in 12-week-old rats. Collagen gels containing PKH26-prelabeled cells were implanted into the defects. In total, 100 µl of normal saline or 500 µM EDA were subcutaneously injected into the defects once daily for the first 4 days. Micro-CT scanning and histological staining were performed to examine bone formation. Immunofluorescence staining for markers of oxidative stress, macrophage, angiogenesis, and osteogenesis was performed.
[Results and Discussion]
EDA suppressed reactive oxygen species production and apoptosis caused by H2O2, recovering cell viability. Decreased expression of osteogenesis-related genes and mineralization caused by H2O2 were restored by EDA. EDA recovered the expression of cell cycle-related genes and enhanced the expression of osteogenesis-related genes. EDA treatment increased new bone volume 2 weeks postoperatively. Fewer CD86-positive M1 and more CD163-positive M2 macrophages were detected in the EDA group. The EDA group also showed stronger immunofluorescence for VEGF and CD31. Additionally, more PKH26-positive and PKH26-osteocalcin-double-positive cells were observed in the EDA group, demonstrating that the survival of the transplanted cells was prolonged and that they differentiated into bone-forming cells (Figure). This could be due to the suppression of oxidative stress at the transplantation site in the EDA group. Local redox control using EDA facilitates bone regeneration by improving the local environment, promoting angiogenesis, prolonging survival, and enhancing the osteogenic capabilities of transplanted cells. Further studies are required to elucidate the mechanisms underlying the effects of EDA on osteogenesis.
[References]
1) Toro-Pérez J, Rodrigo R. Contribution of oxidative stress in the mechanisms of postoperative complications and multiple organ dysfunction syndrome. Redox Rep 2021;26:35–44.
[Purpose]
In cell-based bone augmentation, dysfunction and apoptosis of transplanted cells can occur owing to the oxidative stress caused by the overproduction of reactive oxygen species1. Therefore, this study aimed to investigate the effect of controlling oxidative stress on bone regeneration using a free radical scavenger – edaravone (EDA)
[Methods]
Bone marrow-derived cells were collected from 4-week-old rats. The effects of EDA on cell viability and osteogenic differentiation with or without hydroperioxide (H2O2) were evaluated. RNA sequencing was performed to detect differentially expressed genes. Two critical-sized calvaria defects were created in 12-week-old rats. Collagen gels containing PKH26-prelabeled cells were implanted into the defects. In total, 100 µl of normal saline or 500 µM EDA were subcutaneously injected into the defects once daily for the first 4 days. Micro-CT scanning and histological staining were performed to examine bone formation. Immunofluorescence staining for markers of oxidative stress, macrophage, angiogenesis, and osteogenesis was performed.
[Results and Discussion]
EDA suppressed reactive oxygen species production and apoptosis caused by H2O2, recovering cell viability. Decreased expression of osteogenesis-related genes and mineralization caused by H2O2 were restored by EDA. EDA recovered the expression of cell cycle-related genes and enhanced the expression of osteogenesis-related genes. EDA treatment increased new bone volume 2 weeks postoperatively. Fewer CD86-positive M1 and more CD163-positive M2 macrophages were detected in the EDA group. The EDA group also showed stronger immunofluorescence for VEGF and CD31. Additionally, more PKH26-positive and PKH26-osteocalcin-double-positive cells were observed in the EDA group, demonstrating that the survival of the transplanted cells was prolonged and that they differentiated into bone-forming cells (Figure). This could be due to the suppression of oxidative stress at the transplantation site in the EDA group. Local redox control using EDA facilitates bone regeneration by improving the local environment, promoting angiogenesis, prolonging survival, and enhancing the osteogenic capabilities of transplanted cells. Further studies are required to elucidate the mechanisms underlying the effects of EDA on osteogenesis.
[References]
1) Toro-Pérez J, Rodrigo R. Contribution of oxidative stress in the mechanisms of postoperative complications and multiple organ dysfunction syndrome. Redox Rep 2021;26:35–44.