Anti-chronic Pharyngitis Effect Of Total Flavonoids From Turpinia Arguta Seem And Its Mechanism

Objective:Explore the anti-chronic pharyngitis effect and its mechanism on animal and molecular level,to provide scientific data and theoretical basis for the clinical application of jiangxi province’s specialty medicine Turpinia arguta Seem and the development of new drugs for chronic pharyngitis.Methods:Sixty healthy SD rats,weighing 200-300 grams,were devided into male and female groups.Random principle divided into 10 normal groups,the remaining 50 with 2.5% of the concentration of ammonia into the open-throat sword sprayer to the pharynx of rats to spray the model,once in the morning and once in the afternoon,3 presses each time,continuous spray 16 days.In the normal group,The animals were sprayed with normal saline.The operation method was exactly the same as that in the model groups.The rats’ daily appetite,activity and water consumption,pharyngeal mucosa morphology,color and secretion were observed.After successful modeling,chronic pharyngitis rats were randomly divided into 5 groups,namely the model group,the positive drug group(Manyanshuning Yanyan tablets ig 1.125 g / kg),the TFS low dose group(ig 0.208 g / kg),and the TFS medium dose group(ig 0.416 g /kg).),TFS high-dose group(ig 0.832 g / kg),normal control group was given equal volume of normal saline.After 14 days,the rats were treated,the blood was collected from the eyeballs and then centrifuged.The pharyngeal mucosa and its underlying tissues were removed for examination.HE staining was used to observe the pathological changes of rat pharyngeal tissues;the expression levels of IL-1β,IL-6and TNF-α in rat serum were detected by enzyme-linked immunoassay(ELISA);Western blot was used to detect the expression of NF-κBp65 protein and itsinhibitor IκBα protein in rat pharyngeal tissue.Real-time fluorescent quantitative PCR(RT-q PCR)was used to detect the expression of NF-κBm RNA.Results:1.Observation of general behavior of rats: The model rats hair yellowing,the frequency of drinking water and the amount of water are increased,and the amount of oral secretions is increased.After administration of drugs to the stomach treatment,the state is significantly reduced,such as the disappearance of agitation,drinking water volumn norma,oral secretion gradually decreased,the Total Flavonoids from Turpinia arguta Seem group showed a dose-effect relationship.2.Morphological changes of pharyngeal tissue in rats:The mucosal epithelium of the normal group of rats showed that the mucosal epithdnun was mostly unkcratinizcd flal epithelium;the lamina propria was mostly dense connective tissue;the submucosal layer was rich in mixed glands The glandular structure is normal.Most of the mucosal epithelium of chronic pharyngitis rats is keratinized stratified flat epithelium,which is a apical process with obvious hyperplasia.It is accompanied by a large amount of bleeding and dense infiltration of inflammatory cells.The lamina propria is dense connective tissue,with most connective tissue is fibrotic,with bleeding;submucosal glands are hypertrophic,proliferative,and bleeding.After the drug treatment intervention,the lesions in each group have improved to varying degrees.The positive drug group and the Total Flavonoids from Turpinia arguta Seem high-dose group obviously improved,and the mucous membrane of the pharynx basically returned to normal,with minimal infiltration of inflammatory cells.3.IL-1β,IL-6 and TNF-α expression in rat serum: Compared with the normal group,the expression levels of IL-1β,IL-6 and TNF-α in the model group increased significantly(P<0.01).Compared with the model group,the positive group significantly reduced the levels of IL-1β,IL-6,and TNF-α(P<0.01),and the Total Flavonoids from Turpinia arguta Seem group could reduce IL-1β,IL-6,and TNF-α in chronic pharyngitis rats.The secretion of α showed a certain dose-dependent relationship.The medium and high dose groups of Total Flavonoids from Turpinia arguta Seem had very significant differences in the inhibition of IL-1β in chronicpharyngitis rats(P<0.01).The inhibitory effect of high dose group on chronic pharyngitis rats IL-6 was significantly different(P <0.05,P<0.01).4.Expression of NF-κBp65 protein and IκBα protein in rat pharyngeal tissue:Compared with the normal group,the expression level of NF-κBp65 protein in the model group increased significantly,with a significant difference(P<0.01);compared with the model group,Both the positive group and the Total Flavonoids from Turpinia arguta Seem group could reduce the expression of NF-κBp65 protein,and the Total Flavonoids from Turpinia arguta Seem group showed a certain dose-dependent relationship.The expression of NF-κBp65 protein was significantly down-regulated in the positive group and the medium and high-dose groups of Total Flavonoids from Turpinia arguta Seem,and the difference was extremely significant(P<0.01).Compared with the normal group,the expression level of IκBα protein in the model group was decreased significantly,with a significant difference(P<0.01);compared with the model group,the expression of IκBα protein in the positive group and the Total Flavonoids from Turpinia arguta Seem group increased,and Total Flavonoids from Turpinia arguta Seem group showed a certain dose-dependent relationship.There was a significant difference in the expression of IκBα protein between the positive group and the Total Flavonoids from Turpinia arguta Seem group(P<0.05).The Total Flavonoids from Turpinia arguta Seem group had a significant difference(P<0.01).5.NF-κBm RNA expression in rat pharyngeal tissue: Compared with the normal group,the expression of NF-κBm RNA in the model group increased significantly(P<0.01).Compared with the model group,both the positive group and the Total Flavonoids from Turpinia arguta Seem group could reduce the expression of NF-κBm RNA,and the Total Flavonoids from Turpinia arguta Seem group showed a certain dose-dependent relationship.There was a significant difference in the up-regulation of NF-κBm RNA expression between the positive group and the Total Flavonoids from Turpinia arguta Seem group(P<0.05);the Total Flavonoids from Turpinia arguta Seem group had a significant difference(P<0.01).Conclusion:1.Total Flavonoids from Turpinia arguta Seem have a significant effect on chronic pharyngitis rats,which can obviously improve the pathological morphological variation of pharynx tissue in chronic pharyngitis rats and is dose-dependent.2.The mechanism of anti-chronic pharyngitis induced by Total Flavonoids from Turpinia arguta Seem may be related to its inhibition of NF-κB expression in model rats,promotion of IκBα dissociation,and down-regulation of TNF-α,IL-1β,and IL-6secretion.