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Cross-reacting antibacterial auto-antibodies are produced within coronary atherosclerotic plaques of acute coronary syndrome patients.

Canducci F, Saita D, Foglieni C, Piscopiello MR, Chiesa R, Colombo A, Cianflone D, Maseri A, Clementi M, Burioni R - PLoS ONE (2012)

Bottom Line: In vitro fibrocytes obtained by differentiating CD14+ cells isolated from peripheral blood mononuclear cells also interacted with Fab7816, thus supporting the hypothesis of a specific recognition of fibrocytes into the atherosclerotic lesions.Interestingly, the same antibody, cross-reacted with the outer membrane proteins of Proteus mirabilis and Klebsiella pneumoniae (and possibly with homologous proteins of other enterobacteriaceae present in the microbiota).From all the other three libraries, we were able to clone, by immunoaffinity selection, human monoclonal antibodies cross-reacting with bacterial outer membrane proteins and with transgelin.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Microbiology and Virology, Ospedale San Raffaele, Milan, Italy. Canducci.filippo@hsr.it

ABSTRACT
Coronary atherosclerosis, the main condition predisposing to acute myocardial infarction, has an inflammatory component caused by stimuli that are yet unknown. We molecularly investigated the nature of the immune response within human coronary lesion in four coronary plaques obtained by endoluminal atherectomy from four patients. We constructed phage-display libraries containing the IgG1/kappa antibody fragments produced by B-lymphocytes present in each plaque. By immunoaffinity, we selected from these libraries a monoclonal antibody, arbitrarily named Fab7816, able to react both with coronary and carotid atherosclerotic tissue samples. We also demonstrated by confocal microscopy that this monoclonal antibody recognized human transgelin type 1, a cytoskeleton protein involved in atherogenesis, and that it co-localized with fibrocyte-like cells transgelin+, CD68+, CD45+ in human sections of coronary and carotid plaques. In vitro fibrocytes obtained by differentiating CD14+ cells isolated from peripheral blood mononuclear cells also interacted with Fab7816, thus supporting the hypothesis of a specific recognition of fibrocytes into the atherosclerotic lesions. Interestingly, the same antibody, cross-reacted with the outer membrane proteins of Proteus mirabilis and Klebsiella pneumoniae (and possibly with homologous proteins of other enterobacteriaceae present in the microbiota). From all the other three libraries, we were able to clone, by immunoaffinity selection, human monoclonal antibodies cross-reacting with bacterial outer membrane proteins and with transgelin. These findings demonstrated that in human atherosclerotic plaques a local cross-reactive immune response takes place.

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Related in: MedlinePlus

Biopanning selection with combinatorial IgG/k libraries ID-B, ID-C and ID-D.Each library was independently selected by immunoaffinity on purified OmpK36. Results of screening ELISA assays of 30 clones after four rounds of biopanning selection is shown. Sequence analysis of the positively selected clones (O.D.450 nm >0.25 above background) is shown next to each ELISA screening.
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pone-0042283-g009: Biopanning selection with combinatorial IgG/k libraries ID-B, ID-C and ID-D.Each library was independently selected by immunoaffinity on purified OmpK36. Results of screening ELISA assays of 30 clones after four rounds of biopanning selection is shown. Sequence analysis of the positively selected clones (O.D.450 nm >0.25 above background) is shown next to each ELISA screening.

Mentions: To prove that the local production of antibody clones crossreacting with TAGLN and OMPs in atherosclerotic plaque ID-A was not incidental, nor a single lesion-related finding, the biopanning selection was performed with all three additional libraries from three distinct patients on purified OmpK36 (figure 1 and table S2) since Fab7816 reacted poorly on commercial purified TAGLN in ELISA (Figure S6), OmpK36 was preferred for immunoaffinity selection with the other three libraries. After four independent selection rounds with libraries ID-B, ID-C or ID-D on purified OmpK36, biopanning was stopped and 30 single clones for each library where screened in ELISA on purified OmpK36 and sequenced (Figure 9). Several distinct Fabs, bearing different combinations of heavy and light chains, but all able to specifically bind K.pneumoniae OmpK36, were cloned from each library. For each plaque either the most represented among the selected clones or the best reacting Fab at screening was produced and purified. The obtained representative Fabs were named Fab248 (from plaque ID-B), Fab172 (from plaque ID-C) and Fab1630 (from plaque ID-D).


Cross-reacting antibacterial auto-antibodies are produced within coronary atherosclerotic plaques of acute coronary syndrome patients.

Canducci F, Saita D, Foglieni C, Piscopiello MR, Chiesa R, Colombo A, Cianflone D, Maseri A, Clementi M, Burioni R - PLoS ONE (2012)

Biopanning selection with combinatorial IgG/k libraries ID-B, ID-C and ID-D.Each library was independently selected by immunoaffinity on purified OmpK36. Results of screening ELISA assays of 30 clones after four rounds of biopanning selection is shown. Sequence analysis of the positively selected clones (O.D.450 nm >0.25 above background) is shown next to each ELISA screening.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3412836&req=5

pone-0042283-g009: Biopanning selection with combinatorial IgG/k libraries ID-B, ID-C and ID-D.Each library was independently selected by immunoaffinity on purified OmpK36. Results of screening ELISA assays of 30 clones after four rounds of biopanning selection is shown. Sequence analysis of the positively selected clones (O.D.450 nm >0.25 above background) is shown next to each ELISA screening.
Mentions: To prove that the local production of antibody clones crossreacting with TAGLN and OMPs in atherosclerotic plaque ID-A was not incidental, nor a single lesion-related finding, the biopanning selection was performed with all three additional libraries from three distinct patients on purified OmpK36 (figure 1 and table S2) since Fab7816 reacted poorly on commercial purified TAGLN in ELISA (Figure S6), OmpK36 was preferred for immunoaffinity selection with the other three libraries. After four independent selection rounds with libraries ID-B, ID-C or ID-D on purified OmpK36, biopanning was stopped and 30 single clones for each library where screened in ELISA on purified OmpK36 and sequenced (Figure 9). Several distinct Fabs, bearing different combinations of heavy and light chains, but all able to specifically bind K.pneumoniae OmpK36, were cloned from each library. For each plaque either the most represented among the selected clones or the best reacting Fab at screening was produced and purified. The obtained representative Fabs were named Fab248 (from plaque ID-B), Fab172 (from plaque ID-C) and Fab1630 (from plaque ID-D).

Bottom Line: In vitro fibrocytes obtained by differentiating CD14+ cells isolated from peripheral blood mononuclear cells also interacted with Fab7816, thus supporting the hypothesis of a specific recognition of fibrocytes into the atherosclerotic lesions.Interestingly, the same antibody, cross-reacted with the outer membrane proteins of Proteus mirabilis and Klebsiella pneumoniae (and possibly with homologous proteins of other enterobacteriaceae present in the microbiota).From all the other three libraries, we were able to clone, by immunoaffinity selection, human monoclonal antibodies cross-reacting with bacterial outer membrane proteins and with transgelin.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Microbiology and Virology, Ospedale San Raffaele, Milan, Italy. Canducci.filippo@hsr.it

ABSTRACT
Coronary atherosclerosis, the main condition predisposing to acute myocardial infarction, has an inflammatory component caused by stimuli that are yet unknown. We molecularly investigated the nature of the immune response within human coronary lesion in four coronary plaques obtained by endoluminal atherectomy from four patients. We constructed phage-display libraries containing the IgG1/kappa antibody fragments produced by B-lymphocytes present in each plaque. By immunoaffinity, we selected from these libraries a monoclonal antibody, arbitrarily named Fab7816, able to react both with coronary and carotid atherosclerotic tissue samples. We also demonstrated by confocal microscopy that this monoclonal antibody recognized human transgelin type 1, a cytoskeleton protein involved in atherogenesis, and that it co-localized with fibrocyte-like cells transgelin+, CD68+, CD45+ in human sections of coronary and carotid plaques. In vitro fibrocytes obtained by differentiating CD14+ cells isolated from peripheral blood mononuclear cells also interacted with Fab7816, thus supporting the hypothesis of a specific recognition of fibrocytes into the atherosclerotic lesions. Interestingly, the same antibody, cross-reacted with the outer membrane proteins of Proteus mirabilis and Klebsiella pneumoniae (and possibly with homologous proteins of other enterobacteriaceae present in the microbiota). From all the other three libraries, we were able to clone, by immunoaffinity selection, human monoclonal antibodies cross-reacting with bacterial outer membrane proteins and with transgelin. These findings demonstrated that in human atherosclerotic plaques a local cross-reactive immune response takes place.

Show MeSH
Related in: MedlinePlus