| Purpose:1.Summarize the clinical characteristics of the tutor Song Ruohui in treating acute pharyngitis with a view to further promoting it in clinical practice;2.By constructing a mouse model of pharyngeal virus infection,the therapeutic effect of Zheng’s laryngology Xinliang Yangyin method on acute viral pharyngitis was confirmed,and the possible mechanism of immune inflammation was discussed.Method:1.Through learning from teachers,summarize the clinical experience of tutor Song Ruohui in treating acute pharyngitis.2.Through animal experiments,based on NF-KB pathway,explore the immune inflammatory mechanism of Zheng’s Xin Liang Yang Yin method on acute pharyngitis.2.1.Animal experiment: 108 Balb/c female mice with SPF grade were randomly divided into 6 groups with 18 mice in each group.They are normal group of group A,model group of group B,low dose group of Zizheng Dihuang mixture of group C,medium dose group of Zizheng Dihuang mixture of group D,high dose group of Zizheng Dihuang mixture of group E and ribavirin group of group F.Except for the normal group,the other five groups were all made with RSV nose drops(the previous experiment has proved that the model was successful).After the completion of modeling,group C,D and E were treated with Zizheng Dihuang mixture by gavage;Group F was treated with ribavirin granules by gavage;Group A and group B were treated with equal volume of normal saline.After 1,3 and 5 days of treatment in each group,6 mice were randomly selected each time,and the orbital venous blood was extracted,and the serum IL-1 and TNF were detected by ELLSA method-α,IL-10 content;Then the mice were killed,and the nasopharynx mucosa tissues of the mice after 1,3 and 5 days of treatment were taken for HE staining and IHC staining;After 1,3 and 5 days of treatment,the mouse nasopharynx tissue was taken for WB detection of p-NF in the nasopharynx tissue-κ B protein and p-I κB α Protein expression level.2.2.Data processing: The experimental data were analyzed by SPSS27.0or Graphpad Prism8 software,and the measurement data were expressed in X ± s.When the distribution is normal or the variance is homogeneous,use one-way ANOVA for comparison between multiple groups,LSD method for comparison between two groups,otherwise use Duunett’s T3 method;P<0.05 means the difference is statistically significant.Results:1.The tutor Song Ruohui’s treatment of acute pharyngitis embodies the clinical characteristics of the "xinliang yangyin method ",mainly including the following three aspects:(1)Etiology and pathogenesis: Director Song Ruohui believed that "exogenous wind-heat,heat stagnation in the lung and stomach,phlegm-poison steaming,qi stagnation and blood stasis" were the etiology and pathogenesis of acute pharyngitis,and chronic lesion of the throat was an important factor in its acute attack;(2)Treatment principles and methods: according to the etiology and pathogenesis,the principle of treatment is "xinliang yangyin method","pungent" to dispel wind and dispel evil,"cool" to clear heat and cool blood,"nourishing yin" to strengthen the foundation,and also to promote blood circulation and remove blood stasis.(3)Medication characteristics: add or subtract Zizheng Dihuang Decoction,which is the representative prescription of "Xin Liang Yang Yin Method",and apply it according to the symptoms.2.Animal experiment results:(1)General conditions of mice: the mice in the model group showed poor spirits,fatigue,little movement,hair loss,less luster,less food consumption,and more drinking water,while the mice in the normal group did not show the above symptoms;After the drug intervention,compared with the model group,the above symptoms of the mice in the experimental group,such as the spirit,mood and diet state,were better than before.There was no significant change in body weight of mice in each group.(2)HE staining: After 1 day of treatment,the structure of each layer of mucosa in normal group A mice was intact,and no inflammatory cells were found in each layer;Histopathology of the nasopharyngeal mucosa in groups B,C,D,E and F showed that the structure of the epithelial layer of the mucosa was damaged,disorder,keratosis and hyperplasia,inflammatory cells such as neutrophils and mononuclear macrophages gathered,and the hypertrophy and hyperplasia of the acini were seen in the submucosa.After 3 days of treatment,there was no obvious change in group A;In group B(model group),the epithelial layer was still damaged and keratinized,slightly separated from the lamina propria,and still had inflammatory cell aggregation;The keratinization and proliferation of mucosal epithelial cells in the low and medium dose groups of Zizheng Dihuang Mixture(Group C and Group D)were slightly reduced,and inflammatory hyperemia and infiltration of inflammatory cells were still visible in the tissues;The structure of mucosal epithelium in Zizheng Dihuang mixture high-dose group(group E)and ribavirin group(group F)was not significantly damaged,but there was still slight keratinization and hyperplasia,and the infiltration of inflammatory cells was significantly reduced compared with the previous.After 5 days of treatment,there was no significant change in group A;In group B(model group),the structural destruction and proliferation of mucosal epithelium were slightly improved,and a small amount of inflammatory cells were still visible in the submucosa;In the low and medium dose groups of Zizheng Dihuang Mixture(Group C and D),the epithelial structure was still intact,the cell keratinization was significantly reduced,and there were still mild hyperemia and inflammatory cells;Zizheng Dihuang mixture high-dose group(group E)and ribavirin group(group F)showed that the tissue structure of each layer was basically complete,and no obvious inflammatory cells were found.(3)IHC staining: After 1 day of treatment,RSV-F protein antigen was not found in group A of the normal group,and extensive infection of cells from the epithelial layer of the pharyngeal mucosa to the submucosa was detected in other groups B,C,D,E and F.After 3 days of treatment,extensive infection of cells from the pharyngeal epithelium to the submucosa could be detected in the model group,and there was no significant difference between the model group and the model group;Compared with the one day after treatment,C,D,E,F in the experimental groups showed that the infection of pharyngeal submucosal skin cells was improved,among which the infection in the Zizheng Dihuang mixture high-dose group and ribavirin group decreased most significantly.After 5 days of treatment,the model group could detect that there were still a large number of virus infections in the epithelial layer and lamina propria cells of the pharynx;Compared with 3 days after treatment,the infection of pharyngeal epithelial cells in low dose group C and medium dose group D of Zizheng Dihuang Mixture was significantly improved,and the infection in high dose group E and ribavirin group F basically disappeared.(4)ELISA results: Compare the model group B with the blank group A mice,IL-1 and TNF in the serum of group B-α The content of IL-10 was significantly increased and the content of IL-10 was significantly decreased(P<0.01);Compared with group B,the serum levels of IL-1 and TNF in group C,D,E and F were-α The content of IL-10 was significantly decreased,and the content of IL-10 was significantly increased(P<0.01).The difference was statistically significant,among which Group E and Group F had the most significant change,but the difference between Group E and Group F was not statistically significant(P>0.05).(5)WB results:Compared with group A of blank group,p-NF in the nasopharynx tissue of group B mice-κ B protein expression increased significantly,p-I κ B α The protein expression decreased significantly(P;Compared with model group B,p-NF in the nasopharyngeal tissue of Zizheng Dihuang mixture medium dose group D,high dose group E and ribavirin group F-κ The expression of B protein was significantly reduced,p-I κ B α The protein expression increased significantly(P;However,there was no significant difference between E and F groups(P>0.05).Conclusion:1.Song Ruohui,the inheritor of Zheng’s family,has distinctive clinical characteristics in treating acute pharyngitis by using xinliangyangyin method,which can be further promoted in clinical practice;2.Animal experiments have confirmed that the treatment of pharyngeal virus infection with Zheng’s laryngology Xinliang Yangyin method has prominent effect,which may be through inhibiting virus replication,regulating NF-KB signal pathway,and inhibiting I κ B α Degradation and NF-κ B is activated,thus down-regulating the level of pro-inflammatory factors and up-regulating the level of anti-inflammatory factors to exert the anti-inflammatory effect,providing experimental basis for clinical application. |
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