| Objective:To explore the modulation for immune function and its mechanism of electroacupuncture (EA) at different frequencies under the guidance of Traditional Chinese Medicine (TCM), and to provide the theoretical and practical basis for further clarifying and expanding the clinical application of EA.Methods:Through literature study, to understand the relations of immune and acupuncture, acupuncture and acupoints, acupuncture and immune modulation as well as the clinical application of acupuncture in the view of traditional medicine and modern medical; Neuropharmacology, cytobiology, MTT, ELISA, RIA, immunofluorescence and flow cytometry techniques were used to detect the effect of EA at different frequency on the immunological indexes of NK cell activity, phagocyte function, T cell proliferation and cytokines in humans and mice; To confirm the effect of EA on the level of splenic T cell subsets,(3-endorphin and the expression of a7nAChR in immune cells in mice; Applying a7nAChR antibody and the specific a7nAChR antagonist a-Bgt to study the action of a7nAChR on NK cell activity in EA.Results:1. First, literature studyAcupuncture, which has been practiced for thousands of years, is a gem of traditional Chinese medicine. By regulation of qi and blood, balancing Yin and Yang, dredge the meridians, acupuncture plays a role of enhancing human righteousness and strengthen body resistance and eliminating; The mechanisms that acupuncture modulates immune function is to strengthen the body resistance to consolidate the constitution. The acupoints selection focus on the strong immune-related main point; Acupuncture mediated immune modulation has the characters of integrity and bidirection; EA, developed from acupuncture, has the role of immune modulation which is closely related to the neuroendocrine-immune network systems.2. Second, human experiments(1) The effect of EA of different frequencies on the NK cell activity and phagocytosis (phagocytic rate and phagocytic index):In and1d after the last day of EA with2Hz and2/100Hz, NK cell activity was significantly increased (P<0.05); Only in the first day after EA with a frequence of100Hz, NK cell activity was significantly increased (P<0.05). In and1d after the last day of EA with a frequence of2Hz, the phagocytic percentage rate and phagocytic index were significantly enhanced (P<0.01) and maintain the same level; In and1d after last day of EA with frequence of2/100Hz, the phagocytic percentage and phagocytic index were significantly higher (P<0.05, P<0.01) and there is an upward trend (P<0.0l); In and1d after the last day of EA with a frequence of100Hz, the phagocytic percentage were significantly higher (P<0.01, P<0.05), and only in the last day of EA with a frequent of100Hz the phagocytic index significantly increased (P<0.01), and compared with the last day of EA showed a downward trend (P<0.01).(2) The after-effects of2/100Hz EA on NK cell activity, cytokines and phagocytic function:In the1,3d and the last2d of EA treatment, NK cell activity was significantly increased (P<0.01), and In the3rd day after EA restore to the level before the EA treatment; In and1d after last day of EA, IL-2and IL-6were significantly enhanced (P<0.05), and recovered in the2nd day after EA. while IL-4, IL-10, IL-12, IFN-y and TNF-α had no significant changes; in the1,3d and3days after EA, the phagocytic percentage was significantly higher than the EA level (P<0.01) with the same level in each day and phagocytic index were significantly higher, with an upward trend. There was no significant effect on the white blood cell count treated by2/100Hz EA.(3) The effect of2/100Hz EA at different acupoints on NK cell activity and phagocytic index:NK cell activity and phagocytic index did not change significantly when stimulating the acupoint combination such as right Quchi-right Zhongzhu and left Chengshan-left Kunlun; and when stimulating acupoint combination such as Dazhui-right Quchi and left Zusanli-left Sanyinjiao in the last day of and the3rd day after EA treatment, NK cell activity and phagocytic index changed significantly (P<0.01).3. Third, animal experiments(1) The effect of Single EAS with different frequencies on the immune function of normal mice:After one time of EA, The spleen index, spleen NK cell activity and T cell proliferative ability in mice were detected respectively at0,24and48h time points, the result showed that there was no significant change in the2Hz,100Hz and2/100Hz EA.(2) The effect of successive EAS with different frequencies on immune function of normal mice:After three days of successive EAS with frequence of2Hz,100Hz and2/100Hz, the splenic index did not significantly changed; NK cell activity significantly reduced when treated with100Hz EA (P<0.05), and significantly increased when applied with2/100Hz EA (P<0.05), while there was no significant change with2Hz EA on NK cell activity; Splenic T cell proliferative ability was significantly increased when applied with2Hz EA (P<0.01), while did not change significantly to100Hz and2/100Hz EA; IL-6levels in serum was significantly increased when treated with100Hz and2/100Hz EA (P<0.05, P<0.001), but had no significant change to2Hz EA.(3) The effect of successive2/100Hz EAS on serum cytokines:After treated with successive2/100Hz EAS for3days, the IL-6levels in peripheral blood gradually increased and reached a peak on the3rd day, returned to normal levels on the6th day, and showed significant differences compared with control group on the1st and3rd day (P<0.01), but had no significant changes to the content of IL-2, TNF-a and IFN-y.(4) The effect of successive EAS of different frequencies on T cell subsets in CP mice:After three days of successive EAS with frequence of2Hz,100Hz and2/100Hz, blood of CD4+and CD8+did not significantly changed in2Hz,100Hz and2/100Hz; while CD4+/CD8+was significantly increased when treated with2/100Hz EA (P<0.05), but not in2Hz and100Hz.(5) The effect of successive EAS with different frequencies on β-EP in CP mice:Compared with the control group, in the CP group, β-EP significantly reduced in the hypothalamus (P<0.05), and did not change significantly in medulla oblongata and spleen; compared with the CP group, β-EP did not significantly change in hypothalamus, medulla oblongata and spleen in2Hz EA and100Hz EA, as to2/100Hz EA β-EP did not significantly change in hypothalamus, medulla oblongata, but significantly changed in the spleen (P<0.01), and the level of β-EP were significantly higher than2Hz EA and100Hz EA (P<0.05, P<0.01).(6) The localization of α7nAChR in the mouse macrophage cells:The expression of a7nAChR was found on the surface of macrophages in the Intraperitoneal Fluid of mice with immunofluorescence method (a7nAChR antibody).(7) The effect of antibody against a7nAChR on splenic NK cell activity in100Hz EA:After given a7nAChR antibody to mice, compared with100Hz EA group, NK cell activity was significantly enhanced in100Hz EA plus a7nAChR-Ab group (P<0.05).(8) The effect of specific a7nAChR antagonist a-Bgt on splenic NK cell activity in100Hz EA:After given different doses of specific a7nAChR antagonist a-Bgt to mice, compared to100Hz EA group,1.5and3.0μg specific a7nAChR antagonist a-Bgt plus100Hz EA group were significantly enhanced NK cell activity, respectively (P<0.01).Conclusion:1. Acupuncture has integrity and bidirection for strengthen the body resistance to consolidate the constitution and modulation of body function, its efficacy of acupuncture is closely related to the selection of acupoint;2. EA has immune modulation function and the existence of frequency responsiveness and post-effects, its efficacy is closely related to the number of stimuli; 3. The efficacy of EA immune modulation varied with different acupoints combination, ie. acupoint speciality;4. Interaction between β-endorphin and cytokines is likely to mediate immune modulation function of EA;5.a7nAChR may be involved in100Hz EA modulation for immune function. |
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