Objective: To investigate the effect of different frequencies of electroacupuncture (EA) on motor function and expression of autophagy-related proteins microtubule-associated protein light chain 3 (LC3), etc. in spinal cord injury (SCI) rats, so as to reveal its underlying mechanisms and provide a reference for clinical application.
Methods: A total of 100 male SD rats were randomly divided into sham control, model, 2 Hz-EA, 100 Hz-EA and 2 Hz/100 Hz-EA groups(n=20 in each group). EA (2 Hz, 100 Hz or 2 Hz/100 Hz, 0.4-0.5-0.6 mA increased by 0.1 mA every10 min) was applied to "HuatuoJiaji" (EX-HN1, T9 and T11) for 30 min, once a day for 7 days. The rat's hindlimb motor function was assessed by using Basso, Beattie and Bresnahan(BBB) locomotor rating scale, inclined plane test and plantar imprinting test, separately. The histological changes and neuronal apoptosis of the spinal cord tissue were observed by Fluoro-Jade B (FJB) and Nissl staining, respectively. The expression of LC3, Beclin 1 and cleaved Caspase-3 proteins in the spinal cord was detected by Western blot.
Results: After modeling, the BBB scores and the angles of inclined plane on day 1, 3 and 7 were significantly decreased (P<0.001), and the number of FJB positive cells, and the expression levels of Beclin 1 and cleaved Caspase-3 proteins were considerably increased in the model group compared with the sham control group (P<0.001, P<0.05). After EA intervention, the BBB score and the angles of inclined plate on day 3 in the 2 Hz/100 Hz-EA group (rather than in the 2 Hz- and 100 Hz-EA groups), the BBB score and the angles of inclined plate on day 7 in both 2 Hz/100 Hz and 100 Hz-EA groups(rather than in the 2 Hz-EA group), and the expression levels of Beclin 1 and LC3-Ⅱ/Ⅰ in both 2 Hz/100 Hz and 100 Hz-EA groups (rather than in the 2 Hz-EA group) on day 7 were obviously increased (P<0.001, P<0.01, P<0.05), while the number of FJB-positive neurons in the 3 EA groups, and the expression of cleaved Caspase-3 protein in both 2 Hz/100 Hz and 100 Hz-EA groups and the LC3-Ⅱ/Ⅰ in the 2 Hz-EA group were obviously decreased relevant to the model group (P<0.001, P<0.05). The expression of Beclin 1 in the 100 Hz-EA group was obviously decreased relevant to the 2 Hz/100 Hz-EA group (P<0.05) .Nissl staining displayed appearance of cavities and fuzzy shape of Nissl bodies with light coloration in the injured spinal cord of model group, which was milder in both 2 Hz/100 Hz-EA and 100 Hz-EA groups. Plantar imprinting tests showed dragging gait prints in the model group due to disability in movement, and relatively distinct foot imprints in both 2 Hz/100 Hz and 100 Hz-EA groups.
Conclusion: Both 100 Hz and 2 Hz/100 Hz-EA can effectively promote the recovery of hindlimb locomotor function of SCI rats, which may be related to its function in promoting autophagy of damaged nerve cells and in reducing neuronal apoptosis.
Keywords: Apoptosis; Autophagy; Different frequencies; Electroacupuncture; Spinal cord injury.