More than one disease process in chronic sinusitis based on mucin fragmentation patterns and amino Acid analysis

Abstract

Objective. To characterise fragmentation patterns and amino acid composition of MUC2 and MUC5AC in chronic sinusitis. Methods. Antigenic identity of purified sinus mucins was determined by ELISA. Fragmentation patterns of a MUC5AC rich sample mucin were analysed by Sepharose CL-2B gel chromatography. Samples, divided into one MUC2 rich and one MUC5AC rich group, were subjected to sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and their amino acid contents were analysed. Results. Reduction, trypsin digestion, and papain digestion produced progressively smaller mucin species. On SDS-PAGE, digested MUC5AC rich mucin produced four distinct products. Amino acid analysis was characteristic of mucins with high serine, threonine, and proline contents and reduction and proteolysis increased relative proportions of these amino acids. MUC5AC rich mucins contained more protein than MUC2 rich mucins. Conclusion. Sinus mucin fragmentation produced mucin subunits and glycopeptide units of smaller molecular sizes which are likely to have lower viscoelastic properties. Applying this in vivo could alter mucus physical properties and biologic functions. Amino acid contents of MUC2 and MUC5AC mucins are different. This could be contributing to biological properties and functions of sinus mucins. These data suggest that there may be different pathological processes occurring at the cellular level on chronic sinusitis.

Figure 1

Chromatographic profile of the purified fractionated sinus mucin sample (S1). Sepharose CL-2B gel column (125 × 2.5 cm) was eluted by upward flow with 0.2 M sodium chloride containing 0.02% (w/v) sodium azide and flow rate 18 mL/h. Calibration was firstly performed using 1% (w/v) dextran blue solution containing 0.05% (w/v) methyl orange. The glycoprotein content was estimated by the PAS solution assay. V _o and V _t are the void and total volumes, respectively.

Figure 2

Densitometric scans of SDS-PAGE of polymeric and fragmented MUC5AC and MUC2 rich sinus mucins. The x-axis represents the localization of the different PAS-positive peaks relative to the site of application on the stacking gel (1) and the interface between the stacking and migration gel phases (2). The y-axis represents the relative absorbance of the different PAS-positive material (mucins) measured in arbitrary units. For the purpose of comparison of the different electrophoretic patterns, the presented figures were arranged as follows: ((a), (c), (e), and (g)) polymeric, reduced, papain digested, and trypsin digested MUC5AC rich mucins, respectively; ((b), (d), and (f)) polymeric, reduced, and papain digested MUC2 rich mucins, respectively.