Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The VSports app下载. gov means it’s official. Federal government websites often end in . gov or . mil. Before sharing sensitive information, make sure you’re on a federal government site. .

Https

The site is secure. The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely V体育官网. .

Comparative Study
. 2010 Nov;139(5):1753-61.
doi: 10.1053/j.gastro.2010.07.042. Epub 2010 Jul 23.

α-enolase autoantibodies cross-reactive to viral proteins in a mouse model of biliary atresia

Brandy R Lu  1 , Stephen M Brindley, Rebecca M Tucker, Cherie L Lambert, Cara L Mack
Affiliations
Comparative Study

α-enolase autoantibodies cross-reactive to viral proteins in a mouse model of biliary atresia

Brandy R Lu et al. Gastroenterology. 2010 Nov.

Abstract

Background & aims: Biliary atresia (BA) is a neonatal cholangiopathy of unknown etiology. The bile duct injury that occurs in patients with BA might result from a hepatobiliary viral infection followed by an autoimmune response against the bile duct epithelia VSports手机版. We aimed to identify autoantigens recognized by serum antibodies in the Rhesus rotavirus (RRV)-induced mouse model of BA; findings were correlated with BA in humans. .

Methods: Bile duct epithelial proteins were screened for their reactivity with serum antibodies from the mouse model of BA using immunoblot assays V体育安卓版. Unique proteins that reacted with sera antibodies were identified by mass spectrometry and verified using enzyme-linked immunosorbent assay (ELISA) and immunoblot analyses. Candidate autoantibodies in BA patient sera were analyzed by ELISA. .

Results: A bile duct epithelial antigen that reacted strongly with serum immunoglobulin (Ig) G from the mouse model of BA was identified as α-enolase V体育ios版. α-Enolase autoantibody specificity was confirmed by ELISA and immunoblot analyses. Anti-RRV and anti-enolase antibodies cross-reacted with enolase and RRV proteins; we identified regions of sequence homology between RRV and enolase. Serum samples from patients with BA had increased levels of anti-enolase IgM and IgG. .

Conclusions: We have identified autoantibodies against α-enolase in a mouse model of BA (infected with RRV) and in serum samples from patients, indicating a role of humoral autoimmunity in disease pathogenesis VSports最新版本. The cross-reactivity between an anti-enolase antibody and RRV proteins indicates that molecular mimicry might activate humoral autoimmunity in BA patients; further studies are required. .

PubMed Disclaimer

Conflict of interest statement

Conflicts of interest

The authors disclose no conflicts.

Figures (V体育2025版)

Figure 1
Figure 1
Identification of bile duct epithelial cytosolic proteins reactive to IgG within BA sera. (A) Immunofluorescence: Panel 1: The normal mouse cholangiocyte cell line (NMC) was incubated with anti-cytokeratin 19 (CK19)-AF555 (red) and coun-terstained with nuclear Hoechst dye (blue). Panels 2 and 3: Cultured NMCs were incubated with sera from BA or saline (BSS) control mice, followed by anti-IgG-FITC (green). (B) ELISA: Plates were coated with the NMC cytosolic fraction and incubated with sera followed by anti-IgG HRP and substrate. Quantification of serum IgG reactive to NMC proteins is represented by the mean ± standard error of mean optical density. (C) Western blot analysis: The NMC cytosolic fraction was incubated with sera, and IgG reactive to NMC proteins was visualized by chemiluminescence. The IgG from BA sera reacted to unique proteins at 48 kilodaltons and 33 kilodaltons (boxed bands).
Figure 2
Figure 2
Mass spectrometry and peptide fingerprint analysis identified the 48-kilodalton protein band as α-enolase. (A) Protein analysis: Peptide mass of the 48-kilodalton proteins was determined by MALDI-TOF mass spectrometry, entered into the Mascot search engine, and analyzed against the murine genome. Four candidate proteins were identified as significant, with α-enolase demonstrating the highest peptide coverage. (B) α-Enolase peptide coverage within candidate protein: Shown in bold are the peptides from the candidate protein that contribute to 64% of the total peptide coverage of murine α-enolase.
Figure 3
Figure 3
Generation of anti-enolase antibodies within BA mice is dependent on both RRV infection and biliary obstruction. (A) ELISA: Plates were coated with purified enolase and incubated with sera followed by anti-IgG horseradish peroxidase (HRP) and substrate. Standard curves for anti-enolase were obtained and results shown as mean ± standard error of mean. Control mice included BSS; noncholestatic, RRV-infected siblings of BA (RRV infected, without [w/o] BA); mothers of BA; and bile duct ligated, chole-static adult mice. Significant differences in antibody levels were found between each control group and BA (*P < .05). (B) Western blot analysis: The NMC cytosolic fraction or purified enolase was incubated with sera, and IgG reactive to protein was visualized by chemiluminescence. (C) ELISA verification of RRV infection: Plates were coated with RRV and incubated with sera followed by anti-IgG HRP and substrate. Standard curves for anti-RRV were obtained and results shown as mean ± standard error of mean.
Figure 4
Figure 4
Elevated serum anti-enolase IgM and IgG antibodies in BA patients. (A) Representative immunofluorescence: Panel 1: The human cholangiocyte cell line was incubated with anti-cytokeratin 7-Cy3 (CK7) (red) and counterstained with nuclear Hoechst dye (blue). Panels 2 and 3: Cultured cholangiocytes were incubated with sera from BA or control patients, followed by anti-human IgG-FITC (green). (B) Sera from BA infants at the time of diagnosis (left panel, IgM), BA children >1 year of age with their native liver (right panel, IgG), and other disease age-matched controls were tested for anti-enolase antibodies by ELISA. Each dot represents the average anti-enolase antibody level from a single patient, and the bar is the mean value of all subjects within the group.
Figure 5
Figure 5
Anti-RRV and anti-enolase antibodies cross-reactive with both RRV and enolase proteins. Western blot: Purified RRV, ovalbumin (ova), and enolase (1 μg each) were incubated with either purified anti-RRV or anti-enolase antibody, and reactivity was visualized by chemiluminescence. The ~120-kilodalton RRV protein band reactive to anti-enolase antibody was identified by mass spectrometry to contain proteins VP2, VP4, and VP6.
Figure 6
Figure 6
Sequence homology and ribbon diagrams of α-enolase, VP5, and VP8. (A) BLASTp analysis: Murine α-enolase and RRV protein genome were searched for peptide homology. Three significant matches are shown between α-enolase and VP4 (VP4 subunits are VP5 and VP8). Peptides shown in bold represent an exact match, and a “+” sign between the 2 sequences represents conservative amino acid changes. The underlined peptide segment in VP4 is the VP5 subunit that is a known immunogenic region responsible for generating the neutralizing antibody response. Ribbon diagrams: (B) Enolase and (C) VP5/VP8. Homologous amino acids between α-enolase and VP4 (VP5/VP8 subunits) are highlighted in yellow. The underlined VP5 peptide sequence shown in A that is homologous with enolase is highlighted in purple.
See this image and copyright information in PMC

Comment in

  • Viruses and autoantibodies in biliary atresia.
    Selmi C, Vergani D, Mieli-Vergani G. Selmi C, et al. Gastroenterology. 2010 Nov;139(5):1461-4. doi: 10.1053/j.gastro.2010.09.020. Epub 2010 Sep 25. Gastroenterology. 2010. PMID: 20875483 No abstract available.

References

    1. Sokol RJ, Mack C, Narkewicz MR, et al. Pathogenesis and outcome of biliary atresia: current concepts. J Pediatr Gastroenterol Nutr. 2003;37:4–21. - PubMed
    1. Lykavieris P, Chardot C, Sokhn M, et al. Outcome in adulthood of biliary atresia: a study of 63 patients who survived for over 20 years with their native liver. Hepatology. 2005;41:366–371. - PubMed
    1. Mack CL, Tucker RM, Sokol RJ, et al. Biliary atresia is associated with CD4+ Th1 cell-mediated portal tract inflammation. Pediatr Res. 2004;56:79–87. - PMC - PubMed
    1. Davenport M, Gonde C, Redkar R, et al. Immunohistochemistry of the liver and biliary tress in extrahepatic biliary atresia. J Pediatr Surg. 2001;36:1017–1025. - PubMed
    1. Mack CL. The pathogenesis of biliary atresia: evidence for a virus-induced autoimmune disease. Semin Liver Dis. 2007;27:233–242. - V体育2025版 - PMC - PubMed

Publication types

"V体育安卓版" MeSH terms

Substances