Human intestinal mucus proteins isolated by transanal irrigation and proctosigmoidoscopy

Abstract

<p>Human intestinal mucus essentially consists<br />of a network of Mucin2 glycoproteins<br />embedded in many lower molecular<br />weight proteins. This paper contributes to<br />the proteomic study of human intestinal<br />mucus by comparing two sample collection<br />methods (transanal irrigation and brush<br />cytology during proctosigmoidoscopy) and<br />analysis techniques (electrophoresis and<br />digestion in solution). The entire sample<br />collection and treatment process is explained,<br />including protein extraction, digestion and<br />desalination and peptide characterisation<br />using a nanoAcquity UPLC chromatograph<br />coupled to an HDMS spectrometer equipped<br />with a nanoESI source. Collecting mucus via<br />transanal irrigation provided a larger sample<br />volume and protein concentration from a<br />single patient. The proctosigmoidoscopy<br />sample could be analysed via digestion in<br />solution after depleting albumin. The analysis<br />indicates that a simple mucus lysis method<br />can evaluate the electrophoresis and digestion<br />in solution techniques. Studying human<br />intestinal mucus complexes is important<br />because they perform two essential survival<br />functions for humans as the first biochemical<br />and physical defences for the gastrointestinal<br />tract and a habitat for intestinal microbiota,<br />which are primarily hosted in the colon and<br />exceeds the human genetic information and<br />cell number 100- and 10-fold (1).</p>

A Contribution to the Analytical Proteomics of Human Intestinal Mucus: Sampling, Sample Preparation, and Protein Identification Strategies

“…The protein quantification showed concentrations ranging from 0.65 to 1.12 µg/µL [25]. On the other hand, a single session of transanal irrigation with one patient can provide from 0.5 to 5 mL of HIM, with protein concentrations ranging from 20.0 to 22.4 µg/µL [25], after quantification. This concentration is enough for analysis by SDS-PAGE ( Figure 2) or by 2-D PAGE (Figure 3).…”

“…In fact, this amount of material is not sufficient for evaluation by 2-DE, even using staining reagents like colloidal coomassie, which improves the detection of proteins [24]. The protein quantification showed concentrations ranging from 0.65 to 1.12 µg/µL [25]. On the other hand, a single session of transanal irrigation with one patient can provide from 0.5 to 5 mL of HIM, with protein concentrations ranging from 20.0 to 22.4 µg/µL [25], after quantification.…”