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. 2014 Apr 15;7(5):1911-9.
eCollection 2014.

Protective effects of hyperbaric oxygen treatment against spinal cord injury in rats via toll-like receptor 2/nuclear factor-κB signaling

Jiewen Tan  1 , Fang Zhang  2 , Fang Liang  3 , Yong Wang  4 , Zhuo Li  3 , Jing Yang  3 , Xuehua Liu  3
Affiliations

Protective effects of hyperbaric oxygen treatment against spinal cord injury in rats via toll-like receptor 2/nuclear factor-κB signaling

Jiewen Tan et al. Int J Clin Exp Pathol. .

Abstract

Spinal cord injury (SCI) is a serious medical problem with high mortality and disability rates. Hyperbaric oxygen (HBO) treatment is beneficial for neurological recovery after SCI, but the underlying mechanisms await characterization. This study examined whether HBO treatment following SCI in rats exerts a neuroprotective effect through activation of the toll-like receptor (TLR) 2/nuclear factor (NF)-κB signaling pathway. The SC of rats was injured via T10 laminectomy. Experimental animals (n=144) were divided into four groups: sham-operated (SH), SH+HBO, SCI, and SCI+HBO. Each group was subdivided into six subgroups (n=6 per group) that were examined at 12 h, and 1, 2, 3, 7, and 14 days post-injury. Functional recovery in the hind limb was evaluated using the Basso, Beattie, and Bresnahan (BBB) scoring system. The expression of TLR2 and NF-кB was assessed by real-time polymerase chain reaction and Western blotting, while interleukin-1 (IL)-1β and tumor necrosis factor (TNF)-α levels were measured by enzyme-linked immunosorbent assay. TLR2 and NF-кB levels and histological scores were higher in the SCI than in the SH and SH+HBO groups at various time points. HBO treatment decreased TLR2 and NF-кB expression and histological scores as well as IL-1β and TNF-α levels compared to the SCI group at early post-injury stages. In addition, BBB scores were improved in the SCI+HBO relative to the SCI group at 7 and 14 days. HBO treatment may mitigate secondary injury to the SC by inhibiting inflammatory responses induced by TLR2/NF-кB signaling, thereby promoting functional recovery and improving neurological outcome.

Keywords: Hyperbaric oxygen; nuclear factor κB; spinal cord injury; toll-like receptor 2.

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Figures

Figure 1
Figure 1
Effect of HBO on neurological recovery after SCI. The functional recovery of the hindlimb was evaluated and assigned a Basso, Beattie, and Bresnahan (BBB) score at the indicated time points following SCI. HBO, hyperbaric O2; SCI, spinal cord injury; SH, sham-operated. Data are shown as mean ± standard deviation. *P < 0.05.
Figure 2
Figure 2
Effect of hyperbaric O2 (HBO) on spinal cord tissue after spinal cord injury (SCI). Hematoxylin and eosin-stained sections were examined 1 day after surgery in (A) sham-operated controls (SH); (B) SH animals receiving HBO treatment; (C) animals sustaining SCI; and (D) SCI animals receiving HBO treatment. Tissue damage consisting of edema, hemorrhage, and neutrophil infiltration was observed in the SCI group, which was improved by HBO treatment. Magnification: 400×.
Figure 3
Figure 3
Effect of HBO on histological score after SCI. Scores were assigned based on the presence of edema, neutrophil infiltration, and hemorrhage on a scale of 0 to 4 at the indicated time points after SCI. HBO, hyperbaric O2; SCI, spinal cord injury; SH, sham-operated. Data are shown as mean ± standard deviation. **P < 0.01 vs. SH and SH + HBO groups; #P < 0.05, ##P < 0.01 vs. SCI group.
Figure 4
Figure 4
Effect of HBO on toll-like receptor (TLR) 2 protein expression in the spinal cord after SCI. A: A representative immunoblot is shown. Lane 1, SH; lane 2, SH + HBO; lane 3, SCI; lane 4, SCI + HBO. β-actin was used as a loading control. B: TLR2 expression was quantified from immunoblots at the indicated time points. HBO, hyperbaric O2; SCI, spinal cord injury; SH, sham-operated. Data are shown as mean ± standard deviation.*P < 0.05, **P < 0.01 vs. SH and SH + HBO groups; #P < 0.05 vs. SCI group.
Figure 5
Figure 5
Effect of HBO on nuclear factor (NF)-κBp65 protein expression in the spinal cord after SCI. A: A representative immunoblot is shown. Lane 1, SH; lane 2, SH + HBO; lane 3, SCI; lane 4, SCI + HBO. β-actin was used as a loading control. B: NF-κBp65 expression was quantified from immunoblots at the indicated time points. HBO, hyperbaric O2; SCI, spinal cord injury; SH, sham-operated. Data are shown as mean ± standard deviation. *P < 0.05, **P < 0.01 vs. SH and SH + HBO groups; #P < 0.05, ##P < 0.01 vs. SCI group.
Figure 6
Figure 6
Effect of HBO on toll-like receptor (TLR) 2 and nuclear factor (NF)-κBp65 mRNA expression in the spinal cord after SCI. Transcript levels were measured by real-time quantitative PCR and normalized to the expression level of glyceraldehyde 3-phosphate-dehydrogenase. HBO, hyperbaric O2; SCI, spinal cord injury; SH, sham-operated. Data are shown as mean ± standard deviation. *P < 0.05, **P < 0.01 vs. SH and SH + HBO groups; #P < 0.05, ##P < 0.01 vs. SCI group.
Figure 7
Figure 7
Effect of HBO on interleukin (IL)-1β and tumor necrosis factor (TNF)-α expression after SCI. Cytokine levels in the spinal cord were measured by enzyme-linked immunosorbent assay. HBO, hyperbaric O2; SCI, spinal cord injury; SH, sham-operated. Data are shown as mean ± standard deviation. *P < 0.05, **P < 0.01 vs. SH and SH + HBO groups; #P < 0.05 vs. SCI group.
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