Fragmentation Pattern of Mucins in Normal and Diseased Gastric Mucosae: A Glycoprotein Fractionates with Gastric Mucins Purified from Mucosal Scrapings of Cancer and Peptic Ulcer Patients

Abstract

Background/Aims: Gastric cancer, a fatal malignancy, is prevalent in the Western Cape region of South Africa. The aim of this study was a biochemical characterisation of gastric mucins in this disease, compared with gastric ulceration and controls from transplant donors. Methods: Mucins were extracted in a denaturing medium (to prevent endogenous proteolysis) and purified by caesium chloride density gradient ultracentrifugation. Analysis of mucin was by gel filtration, SDS-PAGE and Western blotting methods. Results: All samples of mucin when analysed by gel filtration were found to contain polymeric glycoprotein together with varying amounts of lower-molecular-weight glycoprotein. SDS-PAGE and Western blot analysis showed that diseased stomachs had glycopeptides of a wider range in size and antigenicity with a greater number of smaller fragments immunoreactive to monoclonal antibodies 2–12M1 and 9–13M1. We identified by SDS-PAGE a glycoprotein which co-fractionates in a caesium chloride density gradient with mucins isolated from gastrectomy specimens resected for carcinoma and peptic ulceration and which was absent from mucins of the transplant donor control group. This neuraminidase-sensitive glycoprotein resisted dissociation from mucin during purification in a 3.5 M CsCl density gradient but was partially separable by Sepharose 2B gel chromatography and heat treatment (100°C, 2.0 min) in SDS. Chemical analysis of the glycoprotein by HPLC favours it being an N-linked glycoprotein. Its non-ideal electrophoretic properties make its exact size estimation difficult and we ascribe to it a broad size range of M_r ∼55–65 kD. Conclusion: We conclude that mucins from diseased stomachs were more degraded than those from donors and that the diseased mucosa reproducibly secretes a M_r ∼55–65 kD glycoprotein, the role of which needs to be established.

The Relationship of a Helicobacter heilmannii Infection to the Mucosal Changes in Abattoir and Laboratory Pig Stomach

“…The BDL pig had an ulceration of the glandular area and of the pars oesophagea. On SDS-PAGE the sham-operated animal showed protein bands in the running gel, thus indicating an extensive fragmentation of mucin, 28 while for the BDL pig there was an absence of staining, thus confirming the histologic findings. There findings do not influence the major theme of this study since only two pigs were involved, but the occurrence of spontaneous ulceration of the glandular regions does form the basis of future studies in the pig ulcer model.…”

“…Although previous studies have reported up to 70% polymer in normal human mucus and less in disease, 33 we were unable to repeat this in our studies of human mucin. 15 Furthermore, SDS-PAGE studies (results not shown) showed varying amounts of subunit (500 kDa) 30 from sample to sample, with no lowmolecular-weight fragments similar to those in gastric cancer 28 in humans.…”

Antiviral Activity of Purified Human Breast Milk Mucin

“…Therefore, an extended rod-like structure of a protein core through the presence of high amounts of proline is not required here. The signature amino acids for mucin are serine, threonine and proline [23] which in this study makes up 20.1% of the mucin, a finding previously shown for a fragment of gastric MUC5AC [25] . In summary, the SDS-PAGE band appearance, Western blotting and amino acid analysis confirmed the milk mucin to be MUC1.…”

Inhibition of Human Immunodeficiency Virus Type 1 Activity by Purified Human Breast Milk Mucin (MUC1) in an Inhibition Assay

“…The purified mucin, identified as MUC1, eluted in the void volume of a Sepharose CL-4B gel filtration column, suggesting it was largely in a polymeric and intact form [28] .…”