Electronic Thesis/Dissertation


Comparative Global Protein Profiling of Pediatric Sinonasal Secretions from Healthy and Chronic Rhinosinusitis Patients Open Access

Chronic rhinosinusitis (CRS) is a highly prevalent disease of the upper respiratory tract and is characterized by mucous overproduction and hyperplasia of the submucosal glands in the sinus mucosa. The composition of sinonasal secretions, however, is minimally characterized. In this study, proteomic analyses were performed to identify the protein components, including specific mucin glycoproteins (mucins), in the sinonasal secretions of control and CRS pediatric patients. Secretions were collected at Children's National Medical Center (CNMC) from (a) CRS patients undergoing functional endoscopic sinus surgery and (b) control patients without CRS undergoing surgical procedures un-related to CRS. Secretions were processed for proteomic analysis using established protocols in the CNMC Proteomics Core. Proteins were separated by electrophoresis on SDS-polyacrylamide gels, bands were excised, digested with trypsin, and analyzed using liquid chromatography tandem mass spectrometry (LC-MS/MS). Proteins were identified using the Sequest algorithm in the Bioworks Browser Software against the UniProt database. Fold-change significance was calculated using the QSpec algorithm. Western blot analysis was performed to validate proteomic findings and to determine whether MUC5B, a glandular mucin, was semi-quantitatively increased in CRS sinonasal secretions. Five samples from CRS patients and three from non-CRS patients have been analyzed by proteomic profiling to date. A total of 294 unique proteins were identified across all samples, some of which were common to both CRS and control samples and others that were differentially expressed, including proteins previously reported to be associated with CRS [BP1 fold-containing family A member 1 (BPIFA1), MUC5B, and chitinase-3-like protein 1 (CHI3L1)]. The protein with the most significant negative fold-change (decreased abundance in CRS samples) was retinal dehydrogenase 1 (ALDH1A1, log2 fold-change = -4.39). The protein with the highest significant positive fold-change in CRS samples was BPIFA1 (log2 fold-change = 3.93). Glycoprotein-340, BPIFB1, S100-A9, and serpin A1 were validated by western blot analyses and showed overall increased expression in CRS samples compared to control samples. Proteomic analyses identified MUC5B mucin in all CRS samples and in two control samples. MUC5AC mucin was identified in three CRS samples and two control samples. The relative abundance based on spectral count data of MUC5B compared to MUC5AC in the CRS samples was significantly higher for MUC5B (p < 0.05). Based on western blot analyses, there was no significant increase in the proportion of MUC5B to MUC5AC expression in CRS samples compared to controls, although there was a trend towards a higher ratio of MUC5B to MUC5AC. Data analysis using protein knowledge database tools revealed that proteins showing the most significant expression in CRS samples compared to non-diseased samples were involved in biological processes such as biological regulation, cellular processes, response to stimuli, and immune system response. Proteomic data suggest that MUC5B is significantly more abundant than MUC5AC in CRS patients, likely reflecting the increased glandular hyperplasia in these patients. Further investigation by proteomic analysis and validation by western blot analysis of proteins present in CRS secretions may provide more insight into the pathogenesis of CRS in pediatric patients

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