Fire Needle Acupuncture Regulates Wnt/ERK Multiple Pathways to Promote Neural Stem Cells to Differentiate into Neurons in Rats with Spinal Cord Injury

Abstract

Background & objective: NSCs therapy is considered one of the most potential methods for spinal cord injury (SCI).

Methods: We build the SCI model rats to investigate the therapeutic effect of fire needle acupuncture in improving the locomotor function of SCI rats and its possible mechanism. BBB scale was used for the motor ability of rats. The expression of Nestin, NSE, Gal-C, and GFAP was detected by immunohistochemistry. Wnt, GSK3β, β-catenin, ERK1/2, CyclinD1, and ngn1 were detected by western blot and PCR. The BBB score of both model group (1.20±0.94, 3.12±0.67, 5.34±1.57, 7.12±1.49) and fire needle group (1.70±0.58, 4.50±1.63, 7.53±2.41, 9.24±0.63) gradually increased after SCI. Furthermore, at d10 and d14, the fire needle group showed a significantly high score compared with that in model group at the same time (P<0.05). Fire needle increased Nestin, NSE, and Gal-C expression inhibited GFAP expression after SCI. Also, fire needle could up-regulate Wnt3a, GSK3β, β-catenin, and ngn1, and down-regulate ERK1/2, cyclinD1 gene and protein expression.

Conclusion: In conclusion, fire needle could improve lower limb locomotor function of SCI rats. Also, fire needles could promote endogenous NSCs proliferation differentiating into neurons, and the mechanism might be mediated by promoting the activation of Wnt/β-catenin and inhibiting the overexpression of ERK.

Keywords: ERK; Fire needle acupuncture; Wnt; neural stem cell; neurons; spinal cord injury..

Fig. (1)

Basso, Beattie, and Bresnahan (BBB) scores of rats in each group at different time points postinjury. Notes: #indicates compared with that in the sham operation group, p<0.05. * indicates compared with that in the model group, p<0.05.

Fig. (2)

BrdU double label results of Nestin, NSE, Gal-C, and GFAP expression of rats in each group at different time point after SCI. (A) Nestin expression of rats in each group at different time point after SCI; (B) NSE expression of rats in each group at different time point after SCI; (C) Gal-C expression of rats in each group at different time point after SCI; (D) GFAP expression of rats in each group at different time point after SCI. Images were processed using Image-pro Plus Software to measure the densities of the double-labeled positive cells.

Fig. (3)

Protein Expression of Wnt3a, GSK3β, β-catenin, ERK1, ERK2, cyclinD1, and ngn1 in each group of different time point (cropped blots from different gels). Notes: B: Blank group; S: Sham operation group; M1: Model group at d1; F1:Fire needle group at d1; M3: Model group at d3; F3:Fire needle group at d3; M7: Model group at d7; F7:Fire needle group at d7; M10: Model group at d10; F10:Fire needle group at d10; M14: Model group at d14; F14:Fire needle group at d14. *indicates that the difference between fire needle group and model group at the same time point is statistically significant (P<0.05). # indicates that the difference between a time point and the point before that in the same group is statistically significant (P<0.05). Δ indicates at d1, there is a significant difference among model group, fire needless group, and sham operation group (P<0.05).

Fig. (4)

Gene expression of Wnt3a, GSK3β, β-catenin, ERK1, ERK2, cyclinD1, and ngn1 in each group of different time point. Notes: *indicates that the difference between fire needle group and model group at the same time point is statistically significant (P<0.05). # indicates that the difference between a time point and the point before that in the same group is statistically significant (P<0.05). Δ indicates at d1, there is a significant difference among model group, fire needless group and sham operation group (P<0.05). needle group were significantly higher than those at d1 and d7, respectively, and the levels at d3, d10, and d14 were significantly higher than those in model group (P<0.05, Fig. 4G).