Effects of electroacupuncture pretreatment on M1 polarization of alveolar macrophages in rats with acute lung injury

Objective:To observe the effects of electroacupuncture(EA)pretreatment on M1 polarization of alveolar macrophages(AMs)in rats with acute lung injury(ALI)induced by lipopolysaccharide(LPS),and to explore the potential protective mechanism of EA.Methods:Forty Sprague-Dawley rats were randomly divided into a normal group,a model group,and three groups of EA pretreatment[including a Chize(LU5)group,a Zusanli(ST36)group and a Chize(LU5)plus Zusanli(ST36)group],with eight rats in each group.The model rats of ALI were established by instilling LPS[2 mg/(kg·bw)]into the trachea of rats for 3 h.The rats in each EA pretreatment group were pretreated with EA for 30 min per day at the corresponding bilateral acupoints 6 d before instilling LPS.Three hours after modeling,the pulmonary function of the rats was tested,and the lung tissue was taken to calculate the ratio of lung wet weight to dry weight(W/D).The pathological lung changes and the injury score were observed by hematoxylin-eosin staining.The contents of tumor necrosis factor(TNF)-α,interleukin(IL)-1β,and myeloperoxidase(MPO)in rat's bronchoalveolar lavage fluid(BALF)were detected by enzyme-linked immunosorbent assay.The mRNA and protein expression levels of M1 macrophage markers clusters of differentiation 86(CD86),inducible nitric oxide synthase(iNOS),and its signaling pathway factor Toll-like receptor(TLR)4,and nuclear factor-κB(NF-κB)p65 in the alveoli were detected by fluorescence quantitative polymerase chain reaction and Western blot,respectively.Results:After being induced by LPS,the pulmonary function of the model rats showed that the forced expiratory volume in 0.1 s(FEV0.1),forced expiratory volume in 0.3 s(FEV0.3),and their respective ratios of FEV to forced vital capacity(FVC)(including FEV0.1/FVC and FEV0.3/FVC)were significantly decreased(P<0.01),while the W/D of lung tissue was increased(P<0.01).The score of lung injury was significantly higher(P<0.01).The contents of TNF-α,IL-1β,and MPO in the BALF and the mRNA and protein expression levels of CD86,iNOS,TLR4,and NF-κB p65 in the lung tissue were significantly increased(P<0.01).After EA pretreatment,the FEV0.1,FEV0.3,FEV0.1/FVC,and FEV0.3/FVC were significantly increased,the lung injury score decreased significantly,and the contents of TNF-α,IL-1β,and MPO in the BALF and the expression levels of CD86,iNOS,TLR4,and NF-κB p65 mRNAs and proteins in the alveoli decreased significantly(P<0.05 or P<0.01).Compared with the other two single acupoint groups,the contents of TNF-α,IL-1β,and MPO in the BALF and the expression levels of CD86,iNOS,TLR4,and NF-κB p65 mRNAs in the alveoli in the Chize(LU5)plus Zusanli(ST36)group were significantly lower(P<0.01).Conclusion:EA pretreatment at Chize(LU5)and Zusanli(ST36)can inhibit inflammation and reduce pulmonary injury in ALI rats induced by LPS.The effect of the combination of Chize(LU5)and Zusanli(ST36)is better than that of using these two acupoints separately,and its mechanism may be related to the inhibition of AMs'M1 polarization by down-regulation TLR4/NF-κB signaling pathway.