健脾柔肝息风汤联合针刺对抽动障碍模型大鼠纹状体区细胞自噬及凋亡的影响

Effect of Jianpi Rougan Xifeng Decoction Combined with Acupunctureon Autophagy and Apoptosis in the Striatal Area of ModelRats with Tic Disorder

【目的】探讨健脾柔肝息风汤联合针刺对抽动障碍模型大鼠的治疗作用及其机制。【方法】采用腹腔注射亚氨基二丙腈(IDPN)方法建立抽动障碍大鼠模型。造模成功后第3天将抽动障碍大鼠随机分为模型组、健脾柔肝息风汤组(汤剂组)、针刺组和健脾柔肝息风汤联合针刺组(联合组),另设空白组,每组12只。给予健脾柔肝息风汤灌胃或联合针刺治疗6周后,评价各组大鼠行为学改变,采用苏木素-伊红(HE)染色法观察各组大鼠纹状体病理学变化,末端脱氧核苷酸转移酶介导的dUTP缺口末端标记(TUNEL)法检测各组大鼠纹状体细胞凋亡,透射电镜观察纹状体组织自噬体形态,Western Blot法检测纹状体中凋亡相关蛋白Bcl-2、Bax、cleaved-caspase-3及自噬相关蛋白微管相关蛋白轻链3(LC3)和Beclin1的蛋白表达水平。【结果】与空白组比较,模型组大鼠运动及刻板行为评分显著升高,纹状体细胞损伤严重,纹状体细胞凋亡率及Bax、cleaved-caspase-3蛋白表达水平显著升高(P<0.05),Bcl-2蛋白表达水平显著降低(P<0.05),而自噬水平及LC3和Beclin1蛋白表达水平无显著性差异(P>0.05);与模型组比较,汤剂组、针刺组和联合组大鼠运动及刻板行为评分显著降低,纹状体细胞损伤明显改善,纹状体细胞凋亡率及Bax、cleaved-caspase-3蛋白表达水平显著降低(P<0.05),自噬水平及Bcl-2、LC3和Beclin1蛋白表达水平显著升高(P<0.05);联合组对上述各指标的改善作用优于汤剂组、针刺组(均P<0.05)。【结论】健脾柔肝息风汤联合针刺治疗可改善抽动障碍大鼠抽动行为,其机制与提高纹状体区细胞自噬水平、抑制细胞凋亡有关。

Objective To investigate the therapeutic effects and its mechanism of Jianpi Rougan Xifeng Decoction combined with acupuncture on model rats with tic disorder.Methods The rat model of tic disorder was established by intraperitoneal injection of iminodipropionitrile(IDPN).On the third day after successful modelling,the tic disorder rats were randomly divided into the model group,the Jianpi Rougan Xifeng Decoction group(decoction group),the acupuncture group and the Jianpi Rougan Xifeng Decoction combined with acupuncture group(combination group),and the blank group was additionally set up,with 12 rats in each group.After 6 weeks of decoction gavage or combined acupuncture treatment,the rats in each group were evaluated for behavioral changes,and hematoxylin-eosin(HE)staining was used to observe the pathological changes in the striatum of each group,and terminal-deoxynucleoitidyl transferase mediated Nick end labeling(TUNEL)method was used to detect apoptosis in the striatum of each group.The morphological features of autophagosomes in striatal tissue under transmission electron microscope.The protein expression levels of apoptosis-related proteins Bcl-2,Bax,cleaved-caspase-3 and the autophagy-related proteins microtubule-associated protein light chain 3(LC3)and Beclin1 in the striatum were measured by Western Blot.Results Compared with the blank group,the scores of motor and stereotyped behavior in the model group were significantly increased,the striatal cells were seriously injured,the apoptosis rate and the protein expressions of Bax and cleaved-caspase-3 were significantly increased(P<0.05),and the protein expression level of Bcl-2 was significantly decreased(P<0.05),but there was no significant difference in autophagy and protein expressions of LC3 and Beclin1(P>0.05).Compared with the model group,the scores of motor and stereotyped behavior in the decoction group,acupuncture group and combination group were significantly decreased,the striatal cell injury was significantly improved,the apoptosis rate of striatal cells and the protein expressions of Bax and cleaved-caspase-3 were significantly decreased(P<0.05),while the level of autophagy and the protein expressions of Bcl-2,LC3 and Beclin1 were significantly increased(P<0.05).The improvement effect of the combination group on the above indexes was superior to that of the decoction group and the acupuncture group(all P<0.05).Conclusion The combination of Jianpi Rougan Xifeng Decoction and acupuncture can improve twitching behavior in rats with tic disorder,and the mechanism is related to increasing the level of autophagy and inhibiting apoptosis in the striatal area.