The Effects And Mechanism Of Yinqiao Powder On Upper Respiratory Tract Mucosal Immune

Objective:Research group found that Yinqiao Powder could enhance the upper airway mucosal immunity function of the upper airway immunity dysfunction model mice, which induced by cold stimulation(UAID model). Afterwards, research team confirmed that Yinqiao Powder had protective function to the UAID model mice, which infected with bacteria through the nasal cavity. In this research, our aims were to study the process of the synthesis and secretion of SIg A. And evaluate the effect of Yinqiao Powder in vivo and explain the mechanism of Yinqiao Powder on acute upper respiratory tract infection(AURI).Methods:1. Copy the the UAID model:Tested the content of secretory immunoglobulin A(SIg A) of mice salivary with the method of Enzyme Linked Immunosorbent Assay(ELISA) to evaluate whether the UAID model copied successly. Then, used the technique of Western Blot and Real-Time PCR to detect the expression of protein of polymeric immunoglobulin receptor(p Ig R),immunoglobulin A(Ig A), and the expression of m RNA of p Ig R. In order to provide experimental basis for the further study of the mechanism of Yinqiao Powder on AURI.2. The research on the mechanism of Yinqiao Powder on mucosal immune of upper respiratory tract2.1 Determine the Ig A and related cytokines:Put the mice to death after the UAID model copied. Took the oral and pharyngeal mucosa of animals. Detected the expression of protein of Ig A of the mucosa with western blot technique and cytokine IL-4, IL-6 and TGF-β by the method of ELISA.;2.2 Determine the PIg R and related cytokines:The UAID model copied and the method of tissue extraction were same as 2.1. Then detected the expression of protein of p Ig R with western blot technique and the expression of m RNA of p Ig R with Real-Time PCR technique. The cytokine IFN-γ and TNF-α were tested by the ELISA. To discuss the possible mechanism of of Yinqiao Powder on upper respiratory tract mucosal immune.3.The protective effect of Yinqiao Powder on the UAID model infected with virus3.1 To determin the LD50 of influenza virus in UAID model:Took the influenza virus liquid stored frozen, thawed after 10 times dilution with 6concentration group of saline 10-1~10-6 standby. Replicated the UAID model, then used the aether anesthetize mice lightly and infected the mice with 10-1~10-6 influenza virus liquid through mouse nasal cavity, after that had a observation continuously for 14 days.Recorded the number of the dead and calculated the mortality of the mice. And then calculated the lethal median dose(LD50) according to the method of Reed-Meunch. To determine the infected concentration of the virus in the UAID model.3.2 The effect of Yinqiao Powder on the mortality and the survival time of the UAID model infected with virus:Replicated the UAID model, then used the aether anesthetize mice lightly and infected the mice with the LD50 influenza virus liquid through the mouse nasal cavity,0.05 ml each one, and recorded the number of the dead mice and survival time of all the animal in each group among the 14 days. And then calculated the mortality rate and life prolonging rate to evaluate the survival protective effect of Yinqiao Powder on the UAID model infected with virus.3.3 The effect of Yinqiao Powder on lung index of the UAID model infected with virus :Groups of experimental animals, dosage, the UAID model copied and the method of virus infected were same as 3.2. Put the mice to death on the 6th day after mice infected,after fetching the blood sample of mice, then removed and weighted the whole lung to calculate the lung index and the lung index inhibition rate. Through observating the lung index to evaluate the protective effect of Yinqiao Powder to lung on the UAID model infected with virus.Results:1.Replication and evaluation of the UAID model:The cold stimulation of-20℃ for 15 minutes could reduce the content of SIg A in salivary(P<0.05), also could decrease the protein expression(P<0.05) and the m RNA levels(P<0.05) of p Ig R. But the expression of Ig A were not significantly influenced.The results showed that the the cold stimulation of-20℃ for 15 minutes could inhibit the secretion of SIg A through reducing the expression of p Ig R, and then decreased the mucosal immunity of upper respiratory tract of mice.2. The research on the mechanism of Yinqiao Powder on mucosal immune of upper respiratory tract2.1 the influence of Yinqiao Powder on Ig A and related cytokines:Yinqiao Powder could increase the IL-6 content of the oral and pharyngeal mucosa of the UAID model mice, the difference was significant(P<0.05), while only had the increasing trend on the expression of Ig A and the content of IL-4 and TGF- β, the difference was not significant(P>0.05).2.2 the influence of Yinqiao Powder on pIg R and related cytokines:Yinqiao Powder could increase the mRNA expression(P<0.01)and the protein expression(P<0.05)of p Ig R of the oral and pharyngeal mucosa of the UAID model mice,so did the content of IFN-γand TNF-α, the difference was significant(P<0.001,P<0.001).3. The protective effect of Yinqiao Powder on the UAID model infected with virus3.1 To determin the LD50 of influenza virus in UAID model:The LD50 of influenza virus in this laboratory is 10-3.33, so took the concentration of influenza virus in 10-3.33 dilution as the infection concentration on the cold mice modelfou the follow experiments.3.2 the effect of Yinqiao Powder on the UAID model infected with virus:Yinqiao Powder could significantly reduce the mortality(P<0.05) and prolong the survival time(P<0.05)of the UAID model mice infected with virus(P<0.05), and could inhibit the lung index(P<0.05) of the mice.Conclusion:1. The cold stimulation could inhibit the secretion of SIg A through reducing the expression level of m RNA and protein of p Ig R, and then decreased the mucosal immunity of upper respiratory tract of mice.2. The mechanism of Yinqiao powder enhancing the mucosal immunity of upper respiratory tract of the UAID model was possibly relate to the prescription could increase the content of SIg A influenced by the up-regulation m RNA expression and protein expression of p Ig R through adding the content of INF-γ and TNF-α of the oral and pharyngeal mucosa of the UAID model mice.3. Yinqiao Powder had protective effect on the UAID model infected with virus. The mechanism of Yinqiao powder on AURI was possibly relate to the prescription could enhancing the mucosal immunity of upper respiratory tract.By increasing content of s Ig A,which influenced by the up-regulation m RNA expression and protein expression of p Ig R through adding the content of INF-γ and TNF-αof the oral and pharyngeal mucosa of the UAID model mice.