Experimentally investigating the effect of buccal acupuncture on analgesic time-effect characteristics and monoamine neurotransmitters

Objective

To investigate the analgesic time-effect characteristics and changes in concentrations of rabbit’s hypothalamic 5-hydroxytryptamine (5-HT) and noradrenaline (NE) caused by buccal acupuncture in the rheumatoid arthritis (RA) rabbits, and to reveal the analgesic central mechanism of buccal acupuncture, thereby providing a theoretical basis for the treatment of pain by buccal acupuncture.

Methods

Forty rabbits were randomly divided into a normal group, a model group, a body acupuncture group, and a buccal acupuncture group, with 10 rabbits in each group. No model was established in the normal group, while equal dose of normal saline was injected at the matched site and time point; rabbits in other groups were subjected to the establishment of RA models using egg protein. From the 27th day of the experiment, rabbits in each group received the designated intervention. Rabbits in the normal group and the model group were fixed for 30 min every day using the same method as those in the other groups. In the acupuncture group, Dubi (ST 35) and Zusanli (ST 36) on bilateral hind limbs were selected. Perpendicular needling (using the needles with 0.25 mm in diameter and 25 mm in length) was performed with twirling manipulation for 15 s at intervals of 5 min. The needles were retained for 30 min and acupuncture was performed once a day. In the buccal acupuncture group, the knee point in the buccal acupuncture and needles with a diameter of 0.25 mm and a length of 15 mm were selected. Oblique needling was performed with twirling manipulation for 15 s at intervals of 5 min. The needles were retained for 30 min and acupuncture was performed once a day. The thermal pain thresholds at the 0, 5, 15, 30, 60, 120 and 240 min after the 1st and 10th acupuncture therapy were measured with a PL-200 thermal-inducing pain meter. After the 10th acupuncture therapy, rabbit’s hypothalamus was removed, and the 5-HT and NE concentrations in the hypothalamus at the peak point of the acupuncture pain threshold curve were determined by high performance liquid chromatography (HPLC).

Results

The analgesic effect was obvious at 5 min after buccal acupuncture started, peaked at 30 min, and decreased to the lowest value at 240 min. Rabbits in the body acupuncture group began to show significant analgesic effect at 15 min, which was peaked at 30 min, and began to decline at 60 min. The pain threshold at 240 min was still higher than that at 0 min. Compared with the model group, the concentrations of hypothalamic 5-HT in the buccal acupuncture group and the body acupuncture group was significantly increased, and the between-group differences were statistically significant (both P<0.05). The NE/5-HT ratios in hypothalamus in the buccal acupuncture group and the body acupuncture group were significantly lower than the ratio in the model group, and the differences were statistically significant (both P<0.05); difference in the decrease was statistically significant between the buccal acupuncture group and the body acupuncture group (P<0.05).

Conclusion

The analgesic effect of buccal acupuncture shows an obvious time-dependent curve. It is characterized by rapid onset of pain relief, rapid increase and decline in pain threshold. 5-HT and NE levels in rabbit’s hypothalamus can be affected by buccal acupuncture, with increased 5-HT concentration and reduced NE/5-HT ratio.

     

基于“肠-关节”轴探讨电针对类风湿关节炎模型家兔肠道菌群及血清炎症因子的影响

目的 观察电针对类风湿关节炎（RA）模型家兔肠道菌群及血清炎性因子的影响,探讨其治疗RA的作用机制。方法 采用卵蛋白诱导联合完全弗氏佐剂建立RA家兔模型,将造模成功家兔随机分为模型组、益生菌组和电针组,每组6只,另取6只家兔作为正常组。电针组电针双侧“足三里”“犊鼻”,30 min/次,益生菌组予益生菌胶囊溶液（14.5 mg/kg）灌胃,均1次/d,连续2周。测量家兔膝关节周径、痛阈,HE染色观察结肠组织和滑膜组织病理形态,16S rDNA测序分析肠道菌群结构变化,ELISA检测血清肿瘤坏死因子-α(TNF-α)、白细胞介素（IL）-1β、IL-6含量。结果 与正常组比较,模型组家兔膝关节周径显著增加、痛阈显著降低（P<0.01）;结肠黏膜损伤严重,杯状细胞缺失,大量炎性细胞浸润;关节囊滑膜表面粗糙,滑膜细胞增生、增厚,炎性细胞浸润明显;肠道菌群物种数目及均匀度减少,变形菌门、厚壁菌门、拟杆菌门相对丰度减少（P<0.05,P<0.01）,脱硫杆菌门相对丰度增加（P<0.01）,弯曲杆菌属、劳森氏菌属及假单胞菌属相对丰度增加（P<0.01）,赫山单胞菌属、...     

颊针对类风湿关节炎家兔镇痛效应特性的研究

目的观察颊针对类风湿关节炎家兔镇痛效应的特点,为颊针临床应用提供实验依据。方法 40只家兔随机分成正常组、模型组、体针组、颊针组,每组10只。采用卵清蛋白诱导关节炎兔模型。造模成功后,体针组家兔在双侧膝眼和足三里、颊针组家兔在颊针"膝"穴每天接受针刺治疗1次,连续10次。分别在治疗第1、4、7、10次测定针刺0、5、15、30、60、120、240 min时的热痛阈值。结果颊针与体针均有明显的镇痛效果。颊针治疗5 min即开始显示明显的镇痛效应,30min时到达峰值,60 min时镇痛效应开始下降,240 min时的痛阈值降到与0 min时相同。而体针在治疗15 min时开始显示明显镇痛效应,30 min时到达峰值,60 min时镇痛效应开始下降,但240min痛阈值仍高于0 min时的痛阈值。结论颊针镇痛效应具有明显的时效关系,其时效曲线与体针时效曲线相似,但颊针起效快于体针,效应持续时间短于体针,且颊针镇痛效应不随治疗次数的增加而变化。     

颊针疗法对类风湿性关节炎兔的镇痛时效及其作用机制

目的：探究针刺对患有类风湿性关节炎大耳白兔的镇痛时效及其中枢作用机制。方法：将60只大耳白兔 随机分为正常组(n=6)、模型组(n=6)、体针组(n=24)和颊针组(n=24),再将后两组分为针后即时(0 h)、针后0.5,1, 2 h 4个亚组,每组各6只。白兔类风湿性关节炎模型采用卵蛋白诱导建立。体针组针刺双侧“膝眼”和“足三里” 1次,颊针组双侧颊针针刺“膝”1次,行针15 s,留针30 min。采用PL-200热刺痛仪检测痛阈,以兔出现缩耳挣扎动 作为疼痛反应指标,自照射开始至兔出现反应的潜伏期时间作为该兔的痛阈值;使用放射免疫法检测兔脑脊液中β- 内啡肽(β-endorplhin,β-EP)及八肽胆囊收缩素(cholecystokinin-8,CCK-8)含量。结果：与正常组相比,模型组痛阈和 CCK-8含量显著降低(P<0.01),β-EP含量显著升高(P<0.05);与模型组相比,体针组和颊针组的痛阈在0和1 h(P<0.05或 P<0.01)显著升高;体针组和颊针组兔脑脊液中CCK-8和β-EP含量均显著高于模型组(P<0.01或P<0.05);颊针组的即时 (0 h)CCK-8和β-EP含量显著高于体针组(P<0.05),其他时间无明显变化。结论：针刺治疗可以有效缓解类风湿性关节 炎的疼痛,颊针疗法的即时镇痛效应优于体针治疗,且这种即时镇痛效应与脑脊液中β-EP和CCK-8含量变化相关。     

热补针法对类风湿关节炎寒证家兔膝关节滑膜组织炎性反应及滑膜细胞坏死性凋亡的影响

目的:观察热补针法对类风湿关节炎(RA)寒证家兔膝关节滑膜组织炎性因子及滑膜细胞坏死性凋亡的影响，探讨热补针法治疗RA的抗炎机制。方法:新西兰家兔按随机数字表法分为正常组、模型组、抑制剂组、捻转补法组、热补针法组，每组8只。采用卵蛋白诱导联合弗氏完全佐剂混合液注射，配合低温冷冻法复制RA寒证模型。抑制剂组予TAK-632(25 mg/kg)灌胃，每2 d治疗1次，共治疗7次；捻转补法组和热补针法组取双侧“足三里”分别施以捻转补法及热补针法操作，均留针30 min, 1次/d,共治疗14次。干预结束后，测量各组家兔膝关节皮温、周径；彩色多普勒超声观察各组家兔膝关节腔积液、滑膜厚度及内部血流信号；HE染色法观察各组家兔膝关节滑膜组织病理形态学变化；ELISA法检测各组家兔血清肿瘤坏死因子(TNF)-α、白细胞介素(IL)-1β及IL-6含量；透射电镜法观察各组家兔膝关节滑膜细胞超微结构及坏死情况；Western blot法检测各组家兔膝关节滑膜组织受体相互作用蛋白(RIPK)1、RIPK3、混合谱系激酶结构域样蛋白(MLKL)、磷酸化(p)-MLKL的表达。结果:与正常组比较，模型组家兔膝...     

基于自噬-NLRP3炎症小体-IL-1β信号轴探究热补针法对类风湿关节炎寒证模型家兔滑膜炎症的影响

目的 观察热补针法对类风湿关节炎（RA）寒证模型家兔滑膜组织自噬-NOD样受体蛋白3(NLRP3)炎症小体-白细胞介素（IL）-1β信号轴的影响，探讨热补针法治疗RA滑膜炎症的作用机制。方法 48只新西兰家兔随机分为正常组、模型组、3-甲基腺嘌呤（3-MA）组、雷帕霉素组、热补针法组及平补平泻组，每组8只。以弗氏完全佐剂+卵蛋白诱导联合低温冷冻法复制RA寒证家兔模型。热补针法组和平补平泻组于“足三里”分别施热补针法及平补平泻针法针刺，留针30 min,1次/d，连续14 d;3-MA组和雷帕霉素组分别耳缘静脉注射3-MA和雷帕霉素溶液，1次/2 d，共7次。免疫组化染色检测膝关节滑膜组织IL-1β表达，免疫荧光染色检测滑膜组织NLRP3表达，透射电镜观察滑膜细胞自噬小体形态，Western blot检测滑膜组织自噬蛋白5(Atg5)、UNC-51样激酶1(ULK1)、微管相关蛋白轻链3(LC3)、Beclin-1蛋白表达。结果 与正常组比较，模型组家兔膝关节周径显著增加、皮温显著降低（P<0.05），滑膜组织IL-1β、NLRP3表达显著升高（P<0.05）,Atg5、UL...     

热补针法对类风湿关节炎寒证家兔滑膜炎性反应及miR-155/TLR4/NF-κB信号轴的影响

目的:观察热补针法对类风湿关节炎(RA)寒证家兔滑膜炎性反应，以及对小核糖核酸-155(miR-155)/Toll样受体4(TLR4)/核转录因子-κB(NF-κB)信号轴的影响，探讨热补针法治疗RA的抑炎机制。方法:将36只家兔随机分为正常组、模型组、激动剂组、抑制剂组、热补针法组(热补组)、激动剂+热补组，每组6只。采用卵蛋白混合弗氏完全佐剂联合低温冷冻法诱导RA寒证家兔模型。激动剂组予腹腔注射miR-155 agomir 4.5 OD;抑制剂组予腹腔注射miR-155 antagomir 6.1 OD;热补组予热补针法针刺双侧“足三里”治疗，留针30 min;激动剂+热补组予腹腔注射miR-155 agomir 4.5 OD,30 min后予热补针法针刺双侧“足三里”治疗，留针30 min。各组干预均1次/d,连续7 d。造模完成后对家兔膝关节行超声检测，测定痛阈和膝关节周径。干预结束后测定家兔痛阈、膝关节周径；采用HE染色法观察家兔膝关节滑膜组织病理形态结构；实时荧光PCR法及Western blot法检测家兔膝关节滑膜组织miR-155、细胞因子信号传导抑制蛋白1(SOCS1...