Experimental Studies On The Relationship Between The Epidermal Growth Factor Receptor Signalling Pathway And Chronic Rhinosinusitis

Experimental Studies on the Relationship between theEpidermal Growth Factor Receptor Signalling Pathwayand Chronic RhinosinusitisPartⅠExperimental Studies on the Relationships betweenMUC5AC and MUC5B and Chronic RhinosinusitisObjective: To investigate the expression of MUC5AC and MUC5B messenger RNAs(mRNAs) and localization of these proteins in human sinus mucosa of chronicrhinosinusitis (CRS) and chronic rhinosinusitis with nasal polyposis (CRS/NP).Methods: Maxillary sinus ostia mucosa was harvested from patients undergoingendoscopic sinus surgery for CRS, CRS/NP, and non-CRS pathologies (control).Sinus mucosa was then analyzed using reverse-transcription polymerase chainreaction (RT-PCR) to detect mRNAs of MUC5AC and MUC5B. Hematoxylin-eosin(HE) staining and immunofluorescent (IF) staining were employed to localizeMUC5AC and MUC5B proteins in the sinus mucosa. Area ratios of positive cells(ARPC) in the epithelia and submucosal glands were compared among the CRS,CRS/NP and normal groups. Results: mRNAs of MUC5AC and MUC5B in the sinusmucosa of CRS and CRS/NP were significantly increased compared to that in normalsinus mucosa (P < 0．01) , and no significant difference was found between the mucosaof CRS and that of CRS/NP (P > 0．05) . Hyperplasia and metaplasia of goblet cells andsubmucosal glands were observed in all cases of CRS and CRS/NP. MUC5AC proteinwas mainly expressed in the goblet cells, while MUC5B expression was located to thesubmucosal glands cells and the epithelia of sinus mucosa. ARPC in staining ofMUC5AC and MUC5B were found no difference between the CRS group and theCRS/NP group (P > 0．05) , whereas significant lower in the normal group compared tothe other two groups, respectively (P < 0．01) . Conclusions: The present studydemonstrated that MUC5AC and MUC5B mucin genes were up-regulated in sinusmucosa of CRS and CRS/NP. MUC5AC and MUC5B may play an important role inthe pathogenesis of chronic rhinosinusitis and nasal polyposis associated withhyperplasia and metaplasia of the goblet cells and submucosal glands.. PartⅡExperimental Studies on the Relationships betweenEGFR and Chronic RhinosinusitisObjective: To investigate the expression of EGFR mRNA in human sinus mucosa andto compare the expression of EGFR and its ligand EGF among a CRS group, aCRS/NP group, and a normal group. Methods: Maxillary sinus ostia mucosa washarvested from patients undergoing endoscopic sinus surgery for CRS, CRS/NP, orfrom patients undergoing surgery for non-CRS pathologies (control). The sampleswere analyzed using RT-PCR to detect mRNA of EGFR. HE staining andimmunofluorescent staining were employed to localize EGFR and EGF proteins in thesinus mucosa. ARPC in the epithelia were compared among the CRS, CRS/NP andnormal groups. Results: The lever of EGFR mRNA in the sinus mucosa of CRS andin that of CRS/NP was significantly increased compared with that in normal sinusmucosa (P < 0．01, respectively), and no significant difference was found between themucosa of CRS and that of CRS/NP (P > 0．05) . EGFR protein was mainly expressedin the goblet cells and basal cells and weakly in the ciliated cells, while EGFexpression was located in epithelial cells and some inflammatory cells but not in thegoblet cells. In the control group, expression of EGFR and EGF proteins was lowercompared to those in CRS and CRS/NP sinus mucosa. No significant ARPCdifferences in staining of EGFR and EGF were found between the CRS group and theCRS/NP group (P > 0．05) , whereas statistically significant differences were foundbetween the normal group and the two CRS groups (P < 0．01) . Conclusion: Theseresults suggest that up-regulation of EGFR cascade may play an important role forMUC production in the sinus mucosa of patients with CRS and CRS/NP.PartⅢStudies for the Roles of Epidermal Growth FactorReceptor Signalling Pathway on Cultured Human NasalEpithelial Cells RPMI-2650Objective: To explore the roles of EGFR signalling pathway on cultured human nasalepithelial cells RPMI-2650．Methods: RPMI-2650 cells were cultured in vitro, andthe cytomorphous was observed using scanning electron microscope. When the cellswere significantly confluent, they were divided into 4 groups: group A: maintained inEMEM medium without adding any stimulators; group B: added with EGF 25ng/ml;group C: added with AG1478 10μM followed by EGF 25ng/ml 30 minutes later; group D: added with PD98O69 30μM followed by EGF 25ng/ml 30 minutes later.After incubated for 24 hours, the expression of EGFR and MUC5AC proteins in thecells of these 4 groups were studied using cytoimmunity and western blotting. Results:RPMI-2650 cells were significantly confluent after incubated for 5 to 7 days. TheEGFR protein was expressed in the cells of group A and D, abundantly in group B,while weakly in group C. The values of comparative optical density (OD) hadsignificant difference between group A,B,D and group C, respectively (P < 0．01) .For the MUC5AC protein, its expression was strong in the cells of group A, abundantin group B, and weak in group C and D. Significant difference of the values ofcomparative OD was analyzed between group B and group C,D, respectively (P <0．01) , while no difference between group C and group D (P > 0．05) . Conclusion: Theproduction of MUC5AC in human nasal epithelial cells RPMI-2650 is regulated viathe expression and activation of epidermal growth factor receptor signalling pathway.