| Acupoint specificity refers to the specificity of acupoints compared with non-acupoints and other acupoints in terms of morphological structure,physiological characteristics,pathological reaction,stimulation effect and other specific characteristics.Acupoint specificity is an important theoretical basis for clinical acupoint selection,and the study on acupoint specificity plays an important role in improving the clinical efficacy of acupoint therapy.Acupoint therapy activate the acupoint effect firstly,which further transmit acupoint stimulation to the target organs and the whole body,so as to achieve the purpose of treating diseases.Hence,the study on the initiation mechanism of acupoint is important for the elucidation of acupoint specificity.Acupoint herbal patching(AHP)has dual therapeutic characteristics of local acupoint effect and pharmacological effect,and is an important part of acupoint therapy.Compared with non-acupoint herbal patching(NAHP),AHP has better therapeutic effects.However,there was a few and fragmented researches on the local effect mechanism of AHP,which limits the further development and clinical application of it.Shenque acupoint,an important acupoint in TCM therapy,is the only acupoint that is visible to the eye and palpable to the hand,and it is closely related to internal organs inside the body.Hence,this paper aims to study the specificity of AHP and its local effect mechanism,using Shenque acupoint as a subject,so as to lay a foundation for further research and clinical application of AHP.1.Study on the specific therapeutic effect of the Shenque acupoint herbal patching(SHP)In this paper,we prepared a rat model of diarrhea by gavage of senna leaves,and carried a compared investigation on the anti-diarrheal effect of SHP with Ding gui’er qi tie(DGE),Xiao’er Fuxie Waifu powder,and cinnamaldehyde gel patch,respectively.The results showed that DGE had the best anti-diarrhea effect,therefore it was used as a model medication in this paper for the following study.This medication was further applied via the Shenque acupoint and non-acupoint of the rats with diarrhea to compare the anti-diarrheal effects of the two therapies.The results showed that compared with NAHP,SHP with DGE had obvious advantages in alleviating diarrhea in rats.2.Study on the transdermal absorption characteristics of drugs via the Shenque acupointIn this paper,the comparation study on percutaneous permeability was carried out to compare the transdermal absorption characteristics of the main active ingredients in the DGE applied on Shenque acupoint and non-acupoint.The results showed that the cumulative permeation amount(Q8)of cinnamaldehyde(CAD),cinnamic acid(CA),and eugenol(EU)through the skin of the Shenque acupoint was higher and the permeation rate(Jss)of them was faster compared with that of the non-acupoint.Whereas the differences in Q8 and Jss of piperine(PIP)through the Shenque acupoint and the non-acupoint skin were not significant.Combined with the existing researches,it was found that the Q8 of drugs with higher molecular weight and log P value through the skin at the Shenque point were quite different from those of the nonacupoints.Since PIP is one of the important active ingredients of DGE,the pharmacokinetic properties of PIP through Shenque acupoint were further investigated in comparison with those of the non-acupoint skin.The results showed that there were two absorption sites for PIP,namely,the dermal microvascular network and subcutaneous vascular.The absorption rate of PIP via the dermal microvascular of the Shenque acupoint skin was slower than that of the nonacupoint,whereas the absorption rate of PIP via the subcutaneous vascular of the Shenque acupoint was faster than that of the non-acupoint.Based on the pharmacokinetic parameters,it was found that PIP showed a slower release pattern via the Shenque acupoint skin compared with the non-acupoint skin.At the meantime,compared with the non-acupoint application,the plasma clearance of PIP through the Shenque acupoint skin was faster and more easily distributed to other parts of the body,increasing its mean retention time,so that the action time of PIP was prolonged.These results showed that compared with the non-acupoint,the absorption characteristics of drugs through the skin at the Shenque point was different,which varied depending on the different physicochemical properties of drugs.3.Study on structural specificity of the Shenque acupoint skinIn this paper,HE staining was applied to compare the skin thickness of the Shenque acupoint and the non-acupoint of rats.The results showed that the stratum corneum(SC)of the Shenque acupoint was significantly thinner than that of the non-acupoint,while there was no significant difference in the thickness of whole skin,viable epidermis and dermis(VE&D)layer of the Shenque acupoint and the non-acupoint.Taking the skin retention amount of CAD,CA,EU and PIP as indicators,Franz diffusion cell method and tape stripping method were used to investigate the reservoir effect of different layers of skin at the Shenque acupoint and that of the non-acupoint on the drugs.The results showed that compared with the non-acupoint,the SC of the Shenque acupoint had a stronger reservoir effect on CAD,and its thickness had no influence on the reservoir effect;the reservoir effect of the Shenque acupoint on CA is weaker than that of the non-acupoint;and the reservoir effect of the SC of the Shenque acupoint on EU and PIP is stronger,but its thickness has an effect on its reservoir effect.Compared with the non-acupoint,the VE&D layers of the Shenque acupoint of rats has the same reservoir effect on the CAD,weaker reservoir effect on CA and EU,and stronger reservoir effect on PIP.These results showed that the reservoir effect of the Shenque acupoint skin to various drugs is different from that of the non-acupoint.Fourier transform infrared spectroscopy was applied to compare the differences of lipids’arrangement and keratin structure of the SC between the Shenque acupoint and the nonacupoint.The results showed that both lipids of the SC of the Shenque acupoint and the nonacupoint were arranged in an orthogonal manner.The proportion of rigid structures in skin keratins of the Shenque acupoint was lower than that of the non-acupoint,while the proportion of random coil structures in skin keratins of the Shenque acupoint was higher.Further studies showed that drugs did not influence the arrangement of lipids in the SC of the Shenque acupoint and the non-acupuncture point,but increased the proportion of random coil structures in skin keratins,especially for the change of keratin structure of the Shenque acupoint.The lipids in SC were sampled by tape stripping method,extracted by Bligh-Dyer method,and detected by LC-MS.The results showed that there was no significant difference in the contents of total ceramide,total free fatty acid and total cholesterol,subclasses of ceramide and cholesterols between the Shenque acupoint and the non-acupoints.However,the content of FA18:8 in the SC of the Shenque acupoint was higher than that of the non-acupoint,while the content of Cer(d18:1/24:1(15Z))and Cer(dl6:1/17:0)in the SC of the Shenque acupoint were lower than those in the SC of the non-acupoint.The amount of tight junction proteins in the skin of rats at the Shenque acupoint and the non-acupoint were determined by Western Blot technique quantitatively.The results showed that the expression of tight junction proteins,containing claudin-1 and occludin,in the skin of the Shenque acupoint were significantly lower than those of the non-acupoint.These results showed that the specifity of skin structure of the Shenque acupoint,including the thinkness of SC,lipid arrangement in the SC,keratin structure,and the amount of tight junction proteins,which were the specific material basis for drugs absorption through the Shenque acupoint skin.4.Study on the regulation effect on skin microcirculation of SHPLaser speckle contrast imaging(LSCI)was applied to compare the effects of SHP with NAHP on local skin microcirculation blood flow of rats.The results showed that DGE could significantly increase the skin microcirculation blood flow perfusion of rats,and the increased amount of skin microcirculation blood flow perfusion in the acupoint area was more than that in the non-acupoint area.Using infrared thermography(IRT)and ELISA method,the regulation mechanism of SHP on skin microcirculation was investigated,with skin surface temperature,concentrations of histamine(HA),5-hydroxytryptamine(5-HT),calcitonin gene-related peptide(CGRP)and substance P(SP)as indexes.The results showed that:① After DGE was applied,the skin surface temperature of the Shenque acupoint and the non-acupoint areas did not increase significantly.② Before application of the medication,the concentrations of HA in the skin of the Shenque acupoint areas of rats were lower than those of the non-acupoint areas,while the concentrations of 5-HT were higher.After application of the medication,the concentrations of HA in the skin of the Shenque acupoint areas of rats increased significantly,but there was no significant change in the skin of the non-acupoint areas,and the concentrations of 5-HT in both areas increased significantly.③ Before administration of the medication,the concentrations of CGRP and SP in the skin of the acupoint areas of rats were higher than those in the non-acupoint areas.After application of the medication,the increases of CGRP and SP in the skin of the acupoint areas of rats were lower than those in the non-acupoint areas.These results showed that the specific effect of SHP on improving the blood flow perfusion of skin microcirculation in the acupoint areas did not depend on the increase of skin surface temperature caused by SHP,but related to the activation of mast cell function and neural responses in the acupoint areas,and the former is the key.After mast cell function was blocked by sodium cromoglicate,LSCI,IRT,and ELISA method were applied to determine skin microcirculation blood perfusion values,skin surface temperature,concentrations of CGRP and SP in the skins of the Shenque acupoint areas of rats,respectively.The results showed that after the blockage of mast cell function,the increase of skin microcirculation perfusion values caused by SHP was also blocked,while the changes of skin surface temperature,the concentrations of CGRP and SP in the skins were not significant,which showed that drug-stimulated mast cell degranulation was the key to improve the blood flow of skin microcirculation in the acupoint area by SHP.5.A preliminary study on the metabolic regulation effect of SHPNon-targeted metabonomics technique was used to compare the changes of metabolites in the Shenque acupoint areas between normal and diseased rats.The results showed that compared with the normal group,immune and inflammatory responses in the acupoint areas of diarrhead rats increased,the metabolic ability of the body decreased,and the syntheses of lipids,proteins and neurotransmitters in the acupoint areas were affected.Furthermore,before and after adminstration of the medication,the changes of metabolites in the Shenque acupoint areas of rats with diarrhea were compared by using non-targeted metabonomics technique.The results showed that SHP could regulate metabolism in the Shenque acupoint areas and further affect the metabolism of the whole body by regulating tricarboxylic acid cycle,sphingolipid metabolism and amino acid metabolism synthesis pathway,so as to achieve the purpose of treating diseases.Finally,the non-targeted metabonomics technique was applied to compare the differences of metabolites between Shenque acupoint and non-acupoint areas of diarrhea rats after administration of DGE.The results showed that from the point of view of local metabolites,SHP had obvious advantages in regulating exercise ability,reducing local inflammatory responces,improving fatigue tolerance and liver metabolism in rats with diarrhea.While the regulation effect on the energy metabolism,the production of steroid hormone and amino acid metabolism of local action areas of SHP were lower than those of NAHP.As above,SHP therapy had obvious advantages over NAHP in the treatment of diarrhea.The mechanisms of local action included:① The skin structure of the acupoint was different from that of the non-acupoint,which made the transdermal absorption of drugs through the skin of the Shenque point showed specific;② The medication regulated vascular tension and increased vascular permeability by promoting the release of HA and 5-HT from mast cells in the acupoint area,so as to increase the blood perfusion of skin microcirculation in the acupoint area,which was helpful for the transdermal absorption of drugs and the transmission of acupoint stimulation;③ SHP therapy had a specific regulatory effect on the metabolites in the acupoint area of rats with diarrhea;these results layed a foundation for the further researches of AHP and the development of acupoint preparations. |
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