针刺百会透曲鬓对脑内出血大鼠脑组织CD32、CD206蛋白表达水平的影响

Effects of needling Baihui(GV20)toward Qubin(GB7)on the protein expression of CD32 and CD206 in the brain tissue of cerebral hemorrhage rats

目的观察针刺百会透曲鬓对脑内出血(intracerebral hemorrhage,ICH)大鼠CD32、CD206蛋白表达水平的影响,探讨其作用机制。方法将60只雄性SD大鼠随机分为A组(假手术组)、B组(模型组)、C组(治疗1组)、D组(治疗2组)和E组(治疗3组),每组12只。每组再随机分造模后1 d和7 d两个亚组,每组6只。采用自体血注入尾壳核法制取ICH模型。造模12 h后,C组采用针刺百会透曲鬓治疗;D组在造模前3 d注射miR-34a-5p agomir-NC,造模12 h后采用针刺百会透曲鬓治疗;E组在造模前3 d注射miR-34a-5p agomir,造模12 h后采用针刺百会透曲鬓治疗。1 d亚组疗程为1 d,7 d亚组疗程为7 d。于相应时间点进行神经功能评分,处死大鼠后收集血肿周围脑组织,使用蛋白质印迹分析和免疫荧光双染检测血肿周围脑组织中CD32、CD206蛋白表达水平、CD32+/Iba-1+和CD206+/Iba-1+双阳性细胞计数变化。结果造模后7 d时,与B组比较,C组ICH大鼠Longa评分降低(P<0.01);与C组比较,E组ICH大鼠Longa评分升高(P<0.05)。造模后1 d、7 d两个时间点,与A组比较,B组ICH大鼠CD32、CD206蛋白表达水平及CD32+/Iba-1+、CD206+/Iba-1+双阳性细胞数量均升高(P<0.01);与B组比较,C组ICH大鼠CD32蛋白表达水平及CD32+/Iba-1+双阳性细胞数量均降低,CD206蛋白表达水平升高,差异均具有统计学意义(P<0.01);与C组和D组比较,E组ICH大鼠CD32蛋白表达水平及CD32+/Iba-1+双阳性细胞数量均升高,CD206蛋白表达水平均降低,差异均具有统计学意义(P<0.01)。造模后7 d时,与B组比较,C组ICH大鼠CD206+/Iba-1+双阳性细胞数量升高(P<0.01);与C组和D组比较,E组ICH大鼠CD206+/Iba-1+双阳性细胞数量均降低,差异均具有统计学意义(P<0.01)。结论针刺百会透曲鬓能抑制ICH大鼠脑组织CD32的蛋白表达,并促进CD206的蛋白表达,可减轻炎性反应,提高机体抗炎能力,缓解神经损伤。

Objective To discuss the effects of needling Baihui(GV20)toward Qubin(GB7)on the protein expression of CD32 and CD206 in the brain tissue of intracerebral hemorrhage(ICH)rats and discuss the mechanism.Method Sixty male Sprague-Dawley(SD)rats were randomized into group A(sham operation group),group B(model group),group C(treatment group 1),group D(treatment group 2),and group E(treatment group 3),with 12 rats in each group.Each group was then further randomized into post-modeling 1 d and 7 d subgroups,each consisting of 6 rats.The ICH model was prepared by autologous blood injection into the caudate putamen.Group C began to receive needling Baihui toward Qubin 12 h after modeling;group D received the injection of miR-34a-5p agomir-NC 3 d before modeling and received needling from Baihui toward Qubin 12 h after modeling;group E received the injection of miR-34a-5p agomir 3 d before modeling and received needling from Baihui toward Qubin 12 h after modeling.The intervention lasted 1 d in the 1 d subgroups,and the intervention lasted 7 d in the 7 d subgroups.The neurologic function was scored at the corresponding time points.The brain tissues around the hematoma were collected after the sacrifice of rats.Western blot analysis and double Immunofluorescence staining were used to detect the protein expression of CD32 and CD206 and double positive cell counts of CD32﹢/Iba-1﹢and CD206﹢/Iba-1﹢in the brain tissue around the hematoma.Result Seven days after modeling,compared with group B,rats in group C had a lower Longa score(P<0.01);compared with group C,group E had a higher Longa score(P<0.05).At the two time points after modeling,1 d and 7 d,compared with group A,rats in group B had higher expression levels of CD32 and CD206 proteins and larger double positive cell counts of CD32﹢/Iba-1﹢and CD206﹢/Iba-1﹢(P<0.01);compared with group B,rats in group C had a lower protein expression level of CD32,smaller double positive cell counts of CD32﹢/Iba-1﹢,and an increased CD206 protein expression level,all showing statistical significance(P<0.01);compared with groups C and D,the protein expression of CD32 and double positive cell counts of CD32﹢/Iba-1﹢increased,and the protein expression level of CD206 dropped in group E,presenting statistical significance(P<0.01).Seven days after modeling,compared with group B,the double positive cell counts of CD206﹢/Iba-1﹢increased in group C(P<0.01);compared with groups C and D,the double positive cell counts of CD206﹢/Iba-1﹢decreased in group E,and the differences were statistically significant(P<0.01).Conclusion Needling from Baihui toward Qubin can inhibit the protein expression of CD32 and promote the protein expression of CD206 in the brain tissue,reduce inflammatory responses,enhance anti-inflammation function,and mitigate neurologic injuries in ICH rats.