Electroacupuncture regulates microglial polarization via inhibiting NF-κB/COX2 pathway following traumatic brain injury

Abstract

BackgroundNeuroinflammation and oxidative stress are important pathological mechanisms following traumatic brain injury (TBI). The NF-κB/COX2 pathway regulates neuroinflammation and oxidative damage, while microglia also play an important role in neuroinflammation. Since NF-κB is involved in microglial polarization, targeting this pathway and microglial polarization is a critical component of TBI treatment. Currently, electroacupuncture (EA) is widely used to treat various symptoms after TBI, but the mechanisms of EA remain poorly understood. Additionally, the optimal frequency of EA remains unclear, which affects its efficacy. This study focuses on exploring the optimal frequency parameters of EA on TBI and investigating the underlying mechanisms of EA through NF-κB/COX2 pathway and microglial polarization. MethodsThe study was divided into two parts. In Experiment 1, 42 Sprague Dawley (SD) rats were induced and randomly divided into seven groups (n = 6). Except for the sham group, all rats underwent controlled cortical impact (CCI) to establish TBI model. Four EA groups (with different frequencies) and manual acupuncture (without current stimulation) received stimulation on the acupoints of Shuigou (GV26), Fengchi (GB20) and Neiguan (PC6) once a day for 7 days. The neurological function was assessed by modified Neurological Severity Scores (mNSS), and the rats’ memory and learning were examined by the Morris water maze (MWM). SOD, MDA, and GSH-Px were detected to evaluate the levels of oxidative stress. The levels of IL-1β, IL-6, and TNF-α were evaluated by Enzyme Linked Immunosorbent Assay (ELISA). Detection of the above indicators indicated a treatment group that exerted the strongest neuroprotection against TBI, we then conducted Experiment 2 using this screened acupuncture treatment to investigate the mechanism of acupuncture. 48 rats were randomly divided into four groups (n = 12): sham, TBI model, acupuncture and PDTC (NF-κB inhibitor). Evaluations of mNSS, MWM test, SOD, MDA, GSH-Px, IL-1β, IL-6, TNF-α, and IL-10 were the same as in Experiment 1. Western blot was applied for detecting the expression levels of NF-κB, p-NF-κB, COX2, and Arg-1. TUNEL was used to examine neuronal apoptosis. Brain structure was observed by H&E. Iba-1, COX2, and Arg-1 were investigated by immunofluorescence staining. ResultsEA with frequency of 2/100 Hz markedly improved neuronal and cognitive function as compared to the other treatment groups. Moreover, it downregulated the expression of MDA, IL-6, IL-1β, and TNF-α and upregulated the levels of SOD and GSH-Px. In addition, Both EA with 2/100 Hz and PDTC reduced the levels of p-NF-κB, COX2 and M1 markers (COX2, IL-6, IL-1β, TNF-α) and increased the levels of M2 markers (Arg-1, IL-10). Moreover, they had similar effects on reducing inflammation, oxidative stress and apoptosis, and improving neuronal and cognitive function. ConclusionsThe collective findings strongly suggest that EA with 2/100 Hz can improve neurologic function by suppressing neuroinflammation, oxidative stress and apoptosis. Additionally, we confirm that EA promotes microglial polarization towards the M2 phenotype through the suppression of NF-κB/COX2 pathway, thus exerting neuroprotective effects after TBI.