The different expressed serum proteins in rhCygb treated rat model of liver fibrosis by the optimized two-dimensional gel electrophoresis.

Liver fibrosis, a common pathological process of chronic liver diseases, is the final stage of liver dysfunction that has severely deleterious impact on human health. Cytoglobin was first discovered in 2001 by proteomic analysis in rat stellate cells and was reported to play an important role in controlling tissue fibrosis. However, the mechanism by which cytoglobin inhibits or reverses the progression of fibrosis remains unclear. The present study examines the effect of recombinant human cytoblobin (rhCygb) in a rat model of liver fibrosis. Proteomic approaches were employed to identify differentially expressed proteins in the fibrosis model. Optimized conditions for two-dimensional gel electrophoresis were developed to provide improved protein detection and separation. A total of 43 spots were obtained and, through the use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry, 30 differentially expressed proteins were identified. Gene ontology term annotation and KEGG pathway analysis allowed us to explore the function of the represented proteins. Based on these results, we provide a theory of the molecular mechanism related to rhCygb reversion of fibrosis and which will assist in the identification of biomarkers in patient serum to improve early diagnosis of liver fibrosis

Anti-hepatic fibrosis effect of rhCygb in CCl₄-induced liver fibrosis model rats( ± s, n = 30).

<p>Anti-hepatic fibrosis effect of rhCygb in CCl₄-induced liver fibrosis model rats( ± s, n = 30).</p

HE and Sirius red staining of liver biopsy samples among the different groups.

<p>A, B and C: the HE staining of liver biopsy of the control group, fibrosis model group and rhCygb treatment group, respectively. D, E and F: the Sirius red staining of liver biopsy samples of the control group, fibrosis model group and rhCygb treatment group, respectively.</p

2-DE analysis of serum samples and the corresponding differentially expressed protein spots.

<p>A: Control group, B: Firbosis model group, C: rhCygb treatment group, D: Fifteen representative differentially expressed protein spots were magnified from three pairs of serum samples. The spots on the left side of the figure were up-regulated in fibrosis model group, while spots on the right were down-regulated in the fibrosis model group. Using a pH3-10, 17cm nonliner IPG Strip and 12.5%SDS-PAGE, we identified 43 different proteins on the gels.</p

Effect after treatment with different kits on 2-DE of serum proteome.

<p>A: Using the Albumin/IgG removal kit, the proteins did not separate well. The main high-abundance proteins were α2-HS glycoprotein, albumin, transferrin, IgA, haptoglobin, Ig heavy chain, α1-antitrypsin, α1-macroglobulin and Ig light chain. B: Use of the ProteinMiner Protein Enrichment Kit resulted in proteins that were separated by a large degree.</p

Analysis of Protein—Protein Interaction Network.

<p>Analysis of Protein—Protein Interaction Network.</p

Distribution of all serum altered proteins into different functional and categories.

<p>a: response to oxidative stress; b: regulation of response to stimulus; c: regulation of immune system process; d: response to inflammatory; e: response to wounding; f: anti-apoptotic process; g: cellular macromolecule catabolic process; h: positive regulation of NF-kB transcription factor activity; i: extracellular region; j: cytosol; k: endosome; l: mitochondrion; m: plasome lipoprotein particle; n: ATP binding; o: ubiquitin protein ligase binding; p: antioxidant activity.</p

GO analysis of the corresponding differentially expressed proteins.

<p>GO analysis of the corresponding differentially expressed proteins.</p