A Study Of Histopathology And Defense Mechanism Of Laryngeal Mucosa In Laryngopharyngeal Reflux Disease

Part 1 Establishment of Laryngopharyngeal Reflux Disease ModelObjective:To establish a model of laryngopharyngeal reflux disease and provide a basis for the study of laryngopharyngeal mucosal histopathological changes in laryngopharyngeal reflux disease.Methods:18 healthy male guinea pigs(300-350g)were randomly divided into 2 groups.10 guinea pigs were perfused with the mixed solution of pepsin and hydrochloride acid(PH=2)for 14 days.The remaining 8 guinea pigs were perfused with normal saline for 14 days.After 14 days,all guinea pigs were sacrificed to obtain histological results.Results: Compared with the normal saline control group,the epithelium of the postcricoid area’s mucosa in the experimental group was partially necrotic and exfoliated,with the infiltration of lamina propria lymphocytes and plasma cells,smooth muscle rupturing,and the hyperplasia of submucosal gland.The leukocyte count of the experimental group was 33.50±16.19.The leukocyte count of normal saline control group was 8.37±3.20.There were statistically significant differences in leukocyte counts between the two groups(P=0.03).Conclusion:The result indicates that acid pepsin can cause inflammation in the throat.Part 2 Dilated Intercelluar Space in the Larynx of Laryngopharyngeal Reflux Disease ModelObjective: To study the changes of the intercellular space of the laryngopharyngeal mucosa epithelium between the experimental group and the control group.Methods: 18 healthy male guinea pigs(300-350g)were randomly divided into 2 groups.10 guinea pigs were perfused with the mixed solution of pepsin and hydrochloride acid(PH=2)for 14 days.The remaining 8 guinea pigs were perfused with normal saline for 14 days.After 14 days,all guinea pigs were sacrificed.The mucosa of the postcricoid area was taken for transmission electron microscopy.Then,we measured the distance of the intercellular space near the basement membrane of the epithelium of the postcricoid area.Results: The distance of the intercellular space of the postcricoid area’s mucosa in the experimental group was 0.32±0.047 um.In the control group,the distance was 0.03±0.014 um.There were statistically significant differences in the distance of intercellular space of the postcricoid area’s mucosa between the two groups(P= 0.000).Conclusion: The result of this study suggests that the morphological change of laryngopharyngeal mucosa in laryngopharyngeal reflux disease is similar to that in gastroesophageal reflux disease and there is widened intercellular space in the laryngeal mucosal epithelium of laryngopharyngeal reflux disease.Part 3 Expression of occludin and claudin-3 in Laryngeal M ucosa of Laryngopharyngeal RefluxObjective: Using the model of laryngopharyngeal reflux to study the expression of occludin and claudin-3 in the laryngopharyngeal mucosa of laryngopharyngeal reflux disease.Methods:18 healthy male guinea pigs(300-350g)were randomly divided into 2 groups.10 guinea pigs were perfused with the mixed solution of pepsin and hydrochloride acid(PH=2)for 14 days.The remaining 8 guinea pigs were perfused with normal saline for 14 days.After 14 days,all guinea pigs were sacrificed to perform immunohistochemical study.Image-pro Plus6.0 was used for analysing the expression differences of o-ccludin and claudin-3 in the mucosa of the postcricoid area of guinea pigs between tow groups.Results: The average optical density of occludin in the experimental group was significantly lower than that in the normal saline control group(P=0.005).The average optical density of claudin-3 in the experimental group was significantly lower than that in the normal saline control group(P=0.009).Conclusion: The expression of occludin and claudin-3 in laryngopharyngeal reflux disease group was significantly down-regulated.It has been confirmed that the impairment of the epithelial barrier in laryngopharyngeal reflux disease is closely related to the tight junction.