| OBJECTIVE:We attempted to investigate the imbalanced states of neuro-endocrine-immuno-modulation through study of the changes of neurikins and their receptors of the tissues of the lung and ileum in experimental rates with spleen-deficiency asthma and simple asthma;to investigate the influence of spleen deficiency on asthma by comparison of the changes of expressive levels between the two kinds of rat models,and thus to provide a new evidence for the view points of "the spleen deficiency as the key patho-mechanism".We attempted the effective mechanism of the therapy of "strengthening the earth and promoting the metal(SEPM)" in treatment of asthma through study of the influence of Chinese herbal remedy of SEPM to the expressive levels of neurikins and their receptors of the tissues of the lung and ileum in experimental rates with spleen-deficiency asthma and simple asthma;to investigate the modern connotation of SEPM theory and the close relationship between the lung and the spleen by studying the correlation between the expressive levels of neurikins and their receptors of the tissues of the lung and ileum in experimental rates with spleen-deficiency asthma and simple asthma.MATERIAL AND METHODS,Taking the theory of traditional Chinese medicine (TCM) as the guide,we set up compound rat models with asthma of spleen deficiency by combination of establishing spleen deficiency animal models in TCM with establishing asthmatic animal models in Western medicine.The spleen models were set up by using the method of improper diet plus physical overstrain.The body weight and temperature of the anus of the animals were compared before and after the model establishment;and the symptoms of the animals were assessed by the standards for evaluation of spleen deficiency models in TCM.The successful animal models were determined by the statistical significance in the changes of their quantitative indexes and their symptomatologic accordance with the standards for evaluation of spleen deficiency.Asthmatic animal models were set up by using ovalbumin as the antigen,and sodium hydroxide and pertusis vaccine as the adjuvant.The asthmatic animals were set up by nebulization following the intraperitoneal injection of valbumin.The successful animal models were evaluated by the appearance of similar symptoms of human asthma in rats such as tachypnea,remarkable motion of nodding,strong construction of abdominal muscle,and occasional pulmonary rale and grunting.The successfully established animal models were given Chinese herbal remedy of SEPM for 12 days.Then the following determinations were conducted:white blood cell count(WBC) and eosinophil count(EOS) in peripheral blood and bronchial alveolar lavage fluid (BALF) by absolute counting;the contents of interleuki-4(IL-4) and interferon-γ(IFN-γ) in BALF by enzyme-linked immunoabsorbent assay(ELISA); the contents of substance P(SP) vasoactive intestinal peptide(VIP) by semi-quantitative analysis of immune-chemical assay;the expressions of SPmRNA and neurokinin 1(NK1) receptor mRNA by reverse transcription-polymerase chain reaction(RT-PCR).RESULTS:1.Compared with the control(C) group,the number of WBC and EOS of the peripheral blood and BALF in the compound(A) group and disease(B) group increased(P＜0.01);the content of IL-4 of BALF in group A and B decreased (P＜0.01),and the content of IFN-γof BALF decreased(P＜0.01),and the ratio of IFN-γ/IL-4 was remarkably decreased(P＜0.01).The mRNA expressions of SPmRNA and NK1 receptor mRNA in group A and B enhanced(P＜0.01);the content of SP in group A and B increased(P＜0.01) and the content of VIP obviously decreased(P＜0.01).2.Compared with group B,the WBC in peripheral blood;the WBC and EOS in BALF;serum IL-4 level,IL-4 level in BALF,the expressions of SPmRNA and NK1 receptor mRNA,and the contents of SP of the lung and ileum tissues in group A all increased remarkably(P＜0.01 or P＜0.05).The level of IFN-γand ratio of IFN-γ/IL-4 in BALF,and the content of VIP in the lung and ileum tissues in group A all decreased remarkably(P＜0.05).3.Compared with the correspondent disease groups,the EOS in blood and BALF in the two Chinese herbal groups(A1,B1) remarkably decreased(P＜0.01 or P＜0.05).And the EOS of BALF in group A1 had showed no significant difference from that in the control group(P＞0.05).The WBC and EOS of blood and BALF in group A1 obviously decreased(P＜0.01);the WBC of BALF in group B1 obviously decreased(P＜0.01).The levels of IL-4 of blood and BALF in both group A1 and B1 remarkably decreased(P＜0.01),but the levels of IFN-γobviously increased (P＜0.01 or P＜0.05);and the ratio of IFN-γ/IL-4 increased(P＜0.01 or P＜0.05).And the level of IFN-γof blood,the level of IL-4 of BALF,and the ratio of IFN-γ/IL-4 of BALF in group A1 had showed no significant difference from those in group C(P＞0.05).The expressions of SPmRNA and NK1 receptor mRNA in group A1 and B1 were remarkably decreased(P＜0.01 or P＜0.05).And the expressive levels of SPmRNA in group A1 and NK1 receptor mRNA in group B1 had showed no significant difference from those in group C(P＞0.05).The contents of SP in the lung and ileum in both group A1 and B1 obviously decreased(P＜0.01),but the content of VIP remarkably increased(P＜0.01).And the contents of SP in the lung and ileum in group A1,and the content of SP in the ileum in group B1 had showed no significant difference from those in group C(P＞0.05).4.There was an obviously negative correlation between the ratio of IFN-γ/IL-4 of blood and the content of SP of the lung tissue in each group(r =-0.698,P＜0.01);there was an obviously positive correlation between the ratio of IFN-γ/IL-4 of blood and the content of VIP of the lung tissue(r=0.741, P＜0.01).There was an obviously negative correlation between the ratio of IFN-γ/IL-4 of blood and the content of SP of the ileum tissue in each group (r=-0.627,P＜0.01);there was an obviously positive correlation between the ratio of IFN-γ/IL-4 of blood and the content of VIP of the ileum tissue(r =0.766,P＜0.01).There was an obviously negative correlation between the ratio of IFN-γ/IL-4 of BALF and the content of SP of the lung tissue in each group (r=-0.688,P＜0.01);there was an obviously positive correlation between the ratio of IFN-γ/IL-4 of BALF and the content of VIP of the lung tissue(r= 0.324,P＜0.01).There was an obviously negative correlation between the ratio of IFN-γ/IL-4 of BALF and the content of SP of the ileum tissue in each group (r=-0.615,P＜0.01);there was an obviously positive correlation between the ratio of IFN-γ/IL-4 of BALF and the content of VIP of the ileum tissue(r= 0.679,P＜0.01).There was an obviously negative correlation between the contents of SP and VIP of the lung tissue in each group(r=-0.678,P＜0.01);there was an obviously positive correlation between the contents of SP and VIP of the lung tissue(r=-0.620,P＜0.01).There was an obviously negative correlation between the contents of SP and VIP of the ileum tissue in each group(r=0.602,P＜0.01);there was an obviously positive correlation between the contents of SP and VIP of the ileum tissue(r=0.608,P＜0.01).5.The pathological results showed that there were bronchial and pulmonary inflammation at different degrees in both group B and A,and the inflammation in both group A1 and B1 had been reduced;there were ileal inflammation at different degrees in both group B and A,and the inflammation in both group A1 and B1 had been reduced,with the inflammation in group A1 was more obvious.CONCLUSION:1.There are,in asthmatic and spleen deficiency asthmatic rats,evident airway or/and general inflammation,expressive imbalance of general and airway immune cytokin IFN-γand IL-4,expressive imbalance of neurikins VIP and SP, and expressive imbalance of the SPmRNA and NK1mRNA receptors of the tissues of the lung and ileum,and the imbalance of neuro-endocrine-immuno-modulation.2.Spleen deficiency can deteriorate the aggregation of inflammatory cells in the peripheral blood and airway of asthmatic rat models,and aggravate the pulmonary and ileal inflammation,and thus exacerbate the general and local inflammatory reaction of the asthmatic body.It can also exacerbate the imbalance of immune cytokins IFN-γand IL-4 in serum and BALF,and aggravate the expressive imbalance of neurikins VIP and SP and the SPmRNA and NK1mRNA receptors of the tissues of the lung and ileum,and thus make the imbalance of neuro-endocrine-immuno-modulation worse,being one of the inner causes for repeated attack of asthma.3.Chinese remedy of SEPM can reduce the airway and general infiltration of inflammatory cells in rat models,down-regulate WBC and EOS in peripheral blood and BALF.It can also regulate the imbalanced state of general and local airway immune cytokins IFN-γand IL-4,up-regulare the content of IFN-γ, down-regulate the content of IL-4,and up-regulate the ratio of IFN-γ/IL-4. It can relieve the expressive imbalance of neurikins VIP,SP and NK1 receptors of the tissues of the lung and ileum,and down-regulate the expressive levels of SPmRNA and NK1mRNA receptors;down-regulate the expressive levels of SP and up-regulate the expressive levels of VIP in both the lung and ileum tissues. And the Chinese remedy of SEPM can reduce the tissue inflammation of the airways, lungs and ileum of the rat models.4.The expressive imbalance of neurikins VIP and SP,and the SPmRNA and NK1mRNA receptors of the tissues of the lung and ileum is obviously correlated with the development and persistence of chronic airway inflammation in asthma, with general and local airway inflammatory state,and with the imbalanced state of general and local airway immune cytokins IFN-γand IL-4.This further reveals the important role of neurikins VIP and SP in development of asthma,especially in inducing imbalance of neuro-endocrine-immuno-modulation.There is evident correlation between the expressive levels of VIP and SP in the tissues of the lung and ileum,which further expounds the close relationship between the lung and the spleen from the angle of neurikins VIP and SP and neuro-endocrine-immuno-modulation. |
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