穴区胶原纤维和肥大细胞在针刺效应中的作用与机制研究
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摘要
大量解剖学和临床医学成像研究已表明人体穴位、经络与结缔组织有着较密切的联系。然而针刺镇痛的效应是如何通过穴位及结缔组织而产生的,至今为止仍然是一个迷。随着技术的发展与研究的进一步深入,各类假说孕育而生。目前,备受关注的一个假说是以Langevin为首的研究组提出的,他们认为针刺疗效的产生可能是由于在行针过程中提插捻转等手法引起了针体周围结缔组织(主要是其中的胶原纤维)的扭曲,从而激发细胞水平的信号循经传导,并形成特异性通路,从而产生疗效。另有报道,穴位处作捻转提插手法后针体上缠绕残留物质主要是一些胶原纤维,这个发现为以上假设提供了有利的证据。至此,有人推测经络的实质可能是人体结缔组织中具有传输信息和能量功能的胶原纤维。
本研究试图通过在不同的针刺条件下,结合形态学观察、力学信号观察及针刺镇痛效应的相关性分析,探索不同针刺方法的穴位始动信号的启动机制。作者首先在离体实验中观察了胶原纤维和肥大细胞在针刺“足三里”穴区筋膜过程中的作用,然后在体实验中,应用急性关节炎(Adjuvant Arthritis,AA)SD大鼠为模型,采用药物穴位注射作为破坏或屏蔽镇痛过程中不同环节的方法,以大鼠后爪痛阈作为针刺效应检测指标,选取常用镇痛穴位“足三里”穴,配合不同针法,采用针刺手法测定仪定量同步记录针体所受机械力学信号。通过所测得的痛阈值,统计出各组痛阈变化率,并辅以穴位组织切片染色及光学显微镜离体穴区胶原纤维与肥大细胞形态变化的观察,比较针刺前后穴位处局部肥大细胞脱颗粒率的变化。
离体实验结果表明:
捻转大鼠“足三里”穴离体筋膜能在短时间内激发毫针附近肥大细胞脱颗粒功能。
在体实验结果表明:
手针(提插、捻转)或电针手法处理后,大鼠炎侧痛阈明显上升,具有良好的镇痛效应(P＜0.05),各组间的镇痛效果无显著性差异(P＞0.05)。
Ⅰ型胶原酶“足三里”穴位预处理破坏胶原纤维后,再施用手针捻转或电针,捻转镇痛效果被抑制(P＞0.05),而电针镇痛效果仍然十分明显(P＜0.05)。色甘酸钠对“足三里”穴位肥大细胞功能进行屏蔽后,再施以手针捻转或电针,也发现手针镇痛效果被抑制(P＞0.05),而电针镇痛效果仍然十分明显(P＜0.05)。
对针刺穴位针体实时受力情况记录发现,“足三里”穴位注射Ⅰ型胶原酶后,再予以捻转或提插,针体提插力或扭矩均有明显下降,较不注射Ⅰ型胶原酶有显著性差异(P＜0.05)。
通过对穴位区组织切片中的肥大细胞的记数和统计,发现手针、电针后肥大细胞脱颗粒率均明显上升,较正常组有显著性差异(P＜0.05)。Ⅰ型胶原酶穴位胶原纤维预处理破坏或色甘酸钠穴区肥大细胞功能屏蔽后,再施用两种手法,肥大细胞脱颗粒率均有下降,较无预处理的情况有显著性差异(P＜0.05)。
“足三里”穴区局部组织切片经胶原纤维特殊染色法染色后显示提插手法组中的胶原纤维表面略显毛糙,捻转手法组中的胶原纤维扭曲并伴有断裂、挛缩现象,Ⅰ型胶原酶穴位预处理后胶原纤维结构模糊,排列较疏散,染色变淡。
“犊鼻”穴Ⅰ型胶原酶预处理后对同经下方“足三里”穴进行提插手针操作,发现针刺镇痛有效(P＜0.05)且“足三里”穴肥大细胞脱颗情况明显,较造模组有显著性差异(P＜0.05)。
研究结果表明胶原纤维参与了手针的针刺始动信号在穴位处的介导过程,通过激活肥大细胞的功能,将有效信息经外周传入到中枢,而电针的针刺信号则是直接激活外周神经感受器由神经介导传入中枢。另外,研究结果提示针刺镇痛效应与肥大细胞脱颗粒率存在着正相关的规律性,同时本实验也为胶原纤维可能是传递针刺信息物质基础的假说提供了证据。
Results from ample anatomy and clinical medical imaging researches indicate that acupuncture points(acupoints) and the structure underlying the meridians have close relationship with connective tissue However,how do the acupuncture analgesic effects act on the aeupoints and the connective tissue remain unknown.Today,the development of the technology and the improvement of the researches bring about all kinds of hypothesizes.Now,there is a famous one suggested by the study group led by Langevin.It is proposed that the deformation of connective tissue during lift-thrust and rotation manipulation at the acupoints,may explain acupuncture therapeutic effects ~([4,5and6].The above mentioned manipulation may act as mechanical stimulus on the connective tissue at the cellular level and may potentially be spread along connective tissue planes.Another report~[7],the residue winded on the needle after being pulled out from the acupoints was mainly collagen fibers~[7],provided powerful evidence to the hypothesis.Hence,it was speculated that the essential material of meridians may be the collagen fibers,whose function is to transmit the information and energy~[8].
In this study,we intend to explore the start-up mechanism for the initiation of acupuncture signal transmission of different acupuncture methods through modality observation,mechanical signal observation and the correlative analysis of acupuncture analgesic effects.The author first observed the role of collagen fibers and mast cells during acupuncturing the fascia of "Zusanli" in vitro experiment.Then in vivo experiment,Arthritis-Acute(AA) were used as rats' model,drugs' injection at acupoints were used to destroy or shield the different parts in acupuncture analgesic effect process and paw withdraw latencies(PWL) of the rats were recorded as the testing indicators of acupuncture effects. Zusanli(ST36),the commonly used acupoint for analgesia studies were chosen as the acupuncture points for different manipulations.Mechanical force,applied to the needle during manipulation,was recorded by a clinical monitor system in real-time.PT change rates were calculated out from the obtained PT data and morphological changes of collagen fibers and mast cells in staining sections of acupoints were observed microscopically.Then the degranulation ratios of mast cells in each group before and after acupuncture were compared.
The experiment results in vitro showed that:
Rotation manipulation could activate the function of mast cells' degranulation around the needle in the fascia of "Zusanli" in short time.
The experiment results in vivo showed that:
Both lift-thrusting and rotation manipulation could raise the PT of AA rats,which had excellent analgesic effects(P＜0.05).However,there was no significant difference between these groups(P＞0.05).
When MA(rotation) was applied after injection of collagenase type-Ⅰin ST36,the analgesic effects were evidently weakened(P＞0.05),but this was not observed for EA(P＜0.05).
Similarly,the effect of MA(rotation) was evidently weakened after injection of DSCG in ST36(P＞0.05),but not observed in EA(P＜0.05).
After analyzing the mechanical data collected during acupuncture,it was found that the force of lift-thrusting or twirling at ST36 declined visibly after collagenase type-Ⅰinjection,and had significant difference compared to those without injection(P＜0.05).
The counting and statistical results of MCs from stained tissue slices showed that the mast cells degranulated obviously after manual and electro acupuncture and had significant difference compared to the group NC(P＜0.05),but the MCs' drgranulation ratios declined if pretreated with type-Ⅰcollagenase or DSCG at "Zusanli" and had significant differences compared to those without pretreatments(P＜0.05). The modality of collagen fibers in stained slices showed that some of the collagen fibers were ruptured after rotation manipulation and some became coarse after lift-thrusting manipulation.After type-Ⅰcollagenase pretreatment,the structure of collagen fibers was destroyed,its arrangement became out of order and the staining color became faded. Lift-thrusting manipulation at Zusanli(ST36) was found of excellent analgesic effects after injection of collagenase type-Ⅰat Dubi(ST35) (P＜0.05),and its MCs degranulation ratios had significant difference compared to group M(P＜0.05).
The results indicated that the collagen fibers participated in the initiation of manual acupuncture signal in the acupoints,and conducted the effective information into centrum by activating the function of MCs. However,the electro-acupuncture signal might be mediated by the peripheral nerve endings directly.In addition,the results suggest that there exists a positive relationship between the MCs degranulation ratios and the acupuncture analgesia and the results also provide evidence to the hypothesis that collagen fiber might be the basic foundation of acupuncture signal conduction.
本研究试图通过在不同的针刺条件下,结合形态学观察、力学信号观察及针刺镇痛效应的相关性分析,探索不同针刺方法的穴位始动信号的启动机制。作者首先在离体实验中观察了胶原纤维和肥大细胞在针刺“足三里”穴区筋膜过程中的作用,然后在体实验中,应用急性关节炎(Adjuvant Arthritis,AA)SD大鼠为模型,采用药物穴位注射作为破坏或屏蔽镇痛过程中不同环节的方法,以大鼠后爪痛阈作为针刺效应检测指标,选取常用镇痛穴位“足三里”穴,配合不同针法,采用针刺手法测定仪定量同步记录针体所受机械力学信号。通过所测得的痛阈值,统计出各组痛阈变化率,并辅以穴位组织切片染色及光学显微镜离体穴区胶原纤维与肥大细胞形态变化的观察,比较针刺前后穴位处局部肥大细胞脱颗粒率的变化。
离体实验结果表明:
捻转大鼠“足三里”穴离体筋膜能在短时间内激发毫针附近肥大细胞脱颗粒功能。
在体实验结果表明:
手针(提插、捻转)或电针手法处理后,大鼠炎侧痛阈明显上升,具有良好的镇痛效应(P＜0.05),各组间的镇痛效果无显著性差异(P＞0.05)。
Ⅰ型胶原酶“足三里”穴位预处理破坏胶原纤维后,再施用手针捻转或电针,捻转镇痛效果被抑制(P＞0.05),而电针镇痛效果仍然十分明显(P＜0.05)。色甘酸钠对“足三里”穴位肥大细胞功能进行屏蔽后,再施以手针捻转或电针,也发现手针镇痛效果被抑制(P＞0.05),而电针镇痛效果仍然十分明显(P＜0.05)。
对针刺穴位针体实时受力情况记录发现,“足三里”穴位注射Ⅰ型胶原酶后,再予以捻转或提插,针体提插力或扭矩均有明显下降,较不注射Ⅰ型胶原酶有显著性差异(P＜0.05)。
通过对穴位区组织切片中的肥大细胞的记数和统计,发现手针、电针后肥大细胞脱颗粒率均明显上升,较正常组有显著性差异(P＜0.05)。Ⅰ型胶原酶穴位胶原纤维预处理破坏或色甘酸钠穴区肥大细胞功能屏蔽后,再施用两种手法,肥大细胞脱颗粒率均有下降,较无预处理的情况有显著性差异(P＜0.05)。
“足三里”穴区局部组织切片经胶原纤维特殊染色法染色后显示提插手法组中的胶原纤维表面略显毛糙,捻转手法组中的胶原纤维扭曲并伴有断裂、挛缩现象,Ⅰ型胶原酶穴位预处理后胶原纤维结构模糊,排列较疏散,染色变淡。
“犊鼻”穴Ⅰ型胶原酶预处理后对同经下方“足三里”穴进行提插手针操作,发现针刺镇痛有效(P＜0.05)且“足三里”穴肥大细胞脱颗情况明显,较造模组有显著性差异(P＜0.05)。
研究结果表明胶原纤维参与了手针的针刺始动信号在穴位处的介导过程,通过激活肥大细胞的功能,将有效信息经外周传入到中枢,而电针的针刺信号则是直接激活外周神经感受器由神经介导传入中枢。另外,研究结果提示针刺镇痛效应与肥大细胞脱颗粒率存在着正相关的规律性,同时本实验也为胶原纤维可能是传递针刺信息物质基础的假说提供了证据。
Results from ample anatomy and clinical medical imaging researches indicate that acupuncture points(acupoints) and the structure underlying the meridians have close relationship with connective tissue However,how do the acupuncture analgesic effects act on the aeupoints and the connective tissue remain unknown.Today,the development of the technology and the improvement of the researches bring about all kinds of hypothesizes.Now,there is a famous one suggested by the study group led by Langevin.It is proposed that the deformation of connective tissue during lift-thrust and rotation manipulation at the acupoints,may explain acupuncture therapeutic effects ~([4,5and6].The above mentioned manipulation may act as mechanical stimulus on the connective tissue at the cellular level and may potentially be spread along connective tissue planes.Another report~[7],the residue winded on the needle after being pulled out from the acupoints was mainly collagen fibers~[7],provided powerful evidence to the hypothesis.Hence,it was speculated that the essential material of meridians may be the collagen fibers,whose function is to transmit the information and energy~[8].
In this study,we intend to explore the start-up mechanism for the initiation of acupuncture signal transmission of different acupuncture methods through modality observation,mechanical signal observation and the correlative analysis of acupuncture analgesic effects.The author first observed the role of collagen fibers and mast cells during acupuncturing the fascia of "Zusanli" in vitro experiment.Then in vivo experiment,Arthritis-Acute(AA) were used as rats' model,drugs' injection at acupoints were used to destroy or shield the different parts in acupuncture analgesic effect process and paw withdraw latencies(PWL) of the rats were recorded as the testing indicators of acupuncture effects. Zusanli(ST36),the commonly used acupoint for analgesia studies were chosen as the acupuncture points for different manipulations.Mechanical force,applied to the needle during manipulation,was recorded by a clinical monitor system in real-time.PT change rates were calculated out from the obtained PT data and morphological changes of collagen fibers and mast cells in staining sections of acupoints were observed microscopically.Then the degranulation ratios of mast cells in each group before and after acupuncture were compared.
The experiment results in vitro showed that:
Rotation manipulation could activate the function of mast cells' degranulation around the needle in the fascia of "Zusanli" in short time.
The experiment results in vivo showed that:
Both lift-thrusting and rotation manipulation could raise the PT of AA rats,which had excellent analgesic effects(P＜0.05).However,there was no significant difference between these groups(P＞0.05).
When MA(rotation) was applied after injection of collagenase type-Ⅰin ST36,the analgesic effects were evidently weakened(P＞0.05),but this was not observed for EA(P＜0.05).
Similarly,the effect of MA(rotation) was evidently weakened after injection of DSCG in ST36(P＞0.05),but not observed in EA(P＜0.05).
After analyzing the mechanical data collected during acupuncture,it was found that the force of lift-thrusting or twirling at ST36 declined visibly after collagenase type-Ⅰinjection,and had significant difference compared to those without injection(P＜0.05).
The counting and statistical results of MCs from stained tissue slices showed that the mast cells degranulated obviously after manual and electro acupuncture and had significant difference compared to the group NC(P＜0.05),but the MCs' drgranulation ratios declined if pretreated with type-Ⅰcollagenase or DSCG at "Zusanli" and had significant differences compared to those without pretreatments(P＜0.05). The modality of collagen fibers in stained slices showed that some of the collagen fibers were ruptured after rotation manipulation and some became coarse after lift-thrusting manipulation.After type-Ⅰcollagenase pretreatment,the structure of collagen fibers was destroyed,its arrangement became out of order and the staining color became faded. Lift-thrusting manipulation at Zusanli(ST36) was found of excellent analgesic effects after injection of collagenase type-Ⅰat Dubi(ST35) (P＜0.05),and its MCs degranulation ratios had significant difference compared to group M(P＜0.05).
The results indicated that the collagen fibers participated in the initiation of manual acupuncture signal in the acupoints,and conducted the effective information into centrum by activating the function of MCs. However,the electro-acupuncture signal might be mediated by the peripheral nerve endings directly.In addition,the results suggest that there exists a positive relationship between the MCs degranulation ratios and the acupuncture analgesia and the results also provide evidence to the hypothesis that collagen fiber might be the basic foundation of acupuncture signal conduction.
引文
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