Korean J Physiol Pharmacol.  2012 Jun;16(3):193-198. 10.4196/kjpp.2012.16.3.193.

Proteomic Analysis of Colonic Mucosal Tissue from Tuberculous and Ulcerative Colitis Patients

• Kwon SC
• Won KJ
• Jung SH
• Lee KP
• Lee DY
• Park ES
• Kim B
• Cheon GJ
• Han KH

• KMID: 1768013
• DOI: http://doi.org/10.4196/kjpp.2012.16.3.193

Abstract

Changes in the expression profiles of specific proteins leads to serious human diseases, including colitis. The proteomic changes related to colitis and the differential expression between tuberculous (TC) and ulcerative colitis (UC) in colon tissue from colitis patients has not been defined. We therefore performed a proteomic analysis of human TC and UC mucosal tissue. Total protein was obtained from the colon mucosal tissue of normal, TC, and UC patients, and resolved by 2-dimensional electrophoresis (2-DE). The results were analyzed with PDQuest using silver staining. We used matrix-assisted laser desorption ionization time-of-flight/time-of-flight spectrometry (MALDI TOF/TOF) to identify proteins differentially expressed in TC and UC. Of the over 1,000 proteins isolated, three in TC tissue and two in UC tissue displayed altered expression when compared to normal tissue. Moreover, two proteins were differentially expressed in a comparative analysis between TC and UC. These were identified as mutant beta-actin, alpha-enolase and Charcot-Leyden crystal protein. In particular, the expression of alpha-enolase was significantly greater in TC compared with normal tissue, but decreased in comparison to UC, implying that alpha-enolase may represent a biomarker for differential diagnosis of TC and UC. This study therefore provides a valuable resource for the molecular and diagnostic analysis of human colitis.

Keyword

Colitis; Tuberculous; Ulcerative; MALDI; Proteomics

MeSH Terms

Actins
Colitis
Colitis, Ulcerative
Colon
Diagnosis, Differential
Electrophoresis
Glycoproteins
Humans
Lysophospholipase
Mucous Membrane
Phosphopyruvate Hydratase
Proteins
Proteomics
Silver Staining
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Spectrum Analysis
Ulcer
Actins
Glycoproteins
Lysophospholipase
Phosphopyruvate Hydratase
Proteins

Figure

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  Fig. 1 Muscosal tissue changes in normal, tuberculous and ulcerative colitis patients. Mucosa tissue was visualized using H&E staining. Arrows and arrow heads in TC show multinucleated giant cells and granuloma with necrosis, UC show crypt abscess, respectively.

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  Fig. 2 Silver-stained 2-DE gels of the colonic proteome of normal, tuberculous and ulcerative colitis patients. Proteins from the colonic mucosa of patients were loaded on IPG pH 3~10 nonlinear (NL) strips and separated by 12% (w/v) polyacrylamide SDS-PAGE. Protein spots were visualized by silver staining. Arrows and numbers correspond to Table 1. 2-DE, 2-dimensional electrophoresis; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; TC, tuberculous colitis; UC, ulcerative colitis.

• 

  Fig. 3 Silver-stained 2-DE images of proteins differentially expressed in tuberculous and ulcerative colitis patients. Enlarged images of proteins found to be differentially expressed by silver-stained 2-DE from Fig. 1. Arrows indicate spots downregulated or upregulated and numbers correspond to the spot numbers shown in Table 1. Graphs show numerical data obtained from images in the left panel. Data were analyzed using the PDQuest software (n=3) *p<0.05 normal tissue vs. TC or UC tissues. TC, tuberculous colitis; UC, ulcerative colitis; OD, optical density.

• 

  Fig. 4 MALDI MS/MS spectra of proteins from the mucosal tissue of colitic patients. Spectra of A (mutant β-actin), B (α-enolase), and C (Charcot-Leyden crystal protein) were obtained using MALDI TOF/TOF.

• 

  Fig. 5 Expression of α-enolase in human TC and UC mucosal tissue. (A) Western blot analysis was performed to determine α-enolalse expression in mucosal tissues from TC and UC patients. (B) Statistical data were obtained from the upper panel. Expression levels in normal tissue are expressed as 100% (n=4). TC, tuberculous colitis; UC, ulcerative colitis.

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