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. 2023 May 4:2023:8227382.
doi: 10.1155/2023/8227382. eCollection 2023.

Hydrogen Promotes the Effectiveness of Bone Mesenchymal Stem Cell Transplantation in Rats with Spinal Cord Injury

Shengchang Luo  1 , Jianxin Wu  2 , Yuanyuan Qiu  3 , Bing Xiao  4 , Yanhai Xi  4 , Chengwei Yang  5 , Zhicai Shi  2 , Weiheng Wang  4
Affiliations

Hydrogen Promotes the Effectiveness of Bone Mesenchymal Stem Cell Transplantation in Rats with Spinal Cord Injury

Shengchang Luo et al. Stem Cells Int. .

Abstract

Although bone mesenchymal stem cell (BMSC) transplantation has been applied to the treatment of spinal cord injury (SCI), the effect is unsatisfactory due to the specific microenvironment (inflammation and oxidative stress) in the SCI area, which leads to the low survival rate of transplanted cells. Thus, additional strategies are required to improve the efficacy of transplanted cells in the treatment of SCI. Hydrogen possesses antioxidant and anti-inflammatory properties. However, whether hydrogen can enhance the effect of BMSC transplantation in the treatment of SCI has not yet been reported. This study was aimed at investigating whether hydrogen promotes the therapeutic effect of BMSC transplantation in the treatment of SCI in rats. In vitro, BMSCs were cultured in a normal medium and a hydrogen-rich medium to study the effect of hydrogen on the proliferation and migration of BMSCs. BMSCs were treated with a serum-deprived medium (SDM), and the effects of hydrogen on the apoptosis of BMSCs were studied. In vivo, BMSCs were injected into the rat model of SCI. Hydrogen-rich saline (5 ml/kg) and saline (5 ml/kg) were given once a day via intraperitoneal injection. Neurological function was evaluated using the Basso, Beattie, and Bresnahan (BBB) and CatWalk gait analyses. Histopathological analysis, oxidative stress, inflammatory factors (TNF-α, IL-1β, and IL-6), and transplanted cell viability were detected at 3 and 28 days after SCI. Hydrogen can significantly enhance BMSC proliferation and migration and tolerance to SDM. Hydrogen and BMSC codelivery can significantly enhance neurological function recovery by improving the transplant cell survival rate and migration. Hydrogen can enhance the migration and proliferation capacity of BMSCs to repair SCI by reducing the inflammatory response and oxidative stress in the injured area. Hydrogen and BMSC codelivery is an effective method to improve BMSC transplantation in the treatment of SCI.

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Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Figure 1
Figure 1
The characterization of BMSCs. (a) BMSCs were observed using a microscope. BMSC-GFP cells showed green fluorescence (b) under a fluorescence microscope. (c–e) The result of BMSCs cultured in osteogenic, lipogenic, and chondrogenic media for 3 weeks. (f) The rates of CD90, CD105, CD73, CD45, CD34, CD11b, and CD19 positivity on P3 BMSCs.
Figure 2
Figure 2
H2 promotes the proliferation ability of BMSCs. (a) The proliferation of BMSCs detected by using EdU staining. (b) Immunofluorescence cell count in EdU detection. (c) Cell proliferation after 0, 6, 12, 24, and 48 h detected by using CCK-8. Data as mean ± SD, ∗P < 0.05 vs. BMSC group (n = 6).
Figure 3
Figure 3
H2 promotes the migration ability of BMSCs. (a) shows the cell migration in the BMSC and H2-BMSC groups at 6 h (73.20 ± 10.43 vs. 158.60 ± 22.11), 12 h (140.60 ± 22.69 vs. 533.20 ± 53.68), and 24 h (761.80 ± 13.27 vs. 747.80 ± 24.03). (b) shows the number of migrated BMSC in the two groups at different times. Data as the mean ± SD. ∗P < 0.05 vs. BMSC group (n = 6).
Figure 4
Figure 4
H2 increases the BMSC tolerance to SDM as detected via flow cytometry. (a) presents the apoptosis rate of BMSCs detected via flow cytometry after tolerance to SDM for 0 and 12 h in the BMSC and H2-BMSC groups. (b) shows the apoptosis rate in the BMSC and H2-BMSC groups. Data as the mean ± SD. ∗P < 0.05 vs. BMSC group (n = 6).
Figure 5
Figure 5
H2 enhances the effect of BMSC transplantation in the treatment of motor function after SCI. (a) shows footprint and walking images of each group 28 days after SCI (blue ink: forelimb prints; red: hindlimb prints). (b–d) show the quantitative analysis of stride length, brake time, and stride time of each group 28 days after SCI. (e) shows the BBB score performed 1 d before surgery and 1 d, 3 d, 7 d, 14 d, 21 d, and 28 d after surgery. Data as mean ± SD. ∗P < 0.05 vs. the sham group. #P < 0.05 vs. the SCI group. &P < 0.05 vs. the BMSC group (n = 10).
Figure 6
Figure 6
HE staining of H2-BMSCs 3 and 28 days after SCI.
Figure 7
Figure 7
H2 enhanced the migration and survival of BMSCs after transplantation. (a) shows green fluorescence of transplanted BMSCs in spinal cord tissue under a fluorescence microscope. (b) shows the count of GFP-positive cells. Data as mean ± SD, ∗P < 0.05 vs. the BMSC group (n = 5).
Figure 8
Figure 8
Determining the content of IL-1β (a), TNF-α (b), and IL-6 (c) via ELISA. Data as mean ± SD. ∗P < 0.05 vs. the sham group. #P < 0.05 vs. the SCI group. &P < 0.05 vs. the BMSC group (n = 5).
Figure 9
Figure 9
Determining the spinal cord oxidative stress of SOD (a) and MDA (b). Data as mean ± SD. ∗P < 0.05 vs. the sham group. #P < 0.05 vs. the SCI group. &P < 0.05 vs. the BMSC group (n = 5).
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