Limits...
Identification of HLA-A2-restricted CD8(+) cytotoxic T cell responses in primary biliary cirrhosis: T cell activation is augmented by immune complexes cross-presented by dendritic cells.

Kita H, Lian ZX, Van de Water J, He XS, Matsumura S, Kaplan M, Luketic V, Coppel RL, Ansari AA, Gershwin ME - J. Exp. Med. (2002)

Bottom Line: Very limited information on autoantigen-specific cytotoxic T lymphocyte (CTL) responses is available compared with autoreactive CD4(+) T cell responses.Furthermore, using soluble PDC-E2 complexed with either PDC-E2-specific human monoclonal antibody or affinity-purified autoantibodies against PDC-E2, the generation of PDC-E2-specific CTLs, occurred at 100-fold and 10-fold less concentration, respectively, compared with soluble antigen alone.The finding that autoantigen-immune complexes can not only cross-present but also that presentation of the autoantigen is of a higher relative efficiency, for the first time defines a unique role for autoantibodies in the pathogenesis of an autoimmune disease.

View Article: PubMed Central - PubMed

Affiliation: Division of Rheumatology, Allergy and Clinical Immunology, University of California at Davis School of Medicine, Davis, CA 95616, USA.

ABSTRACT
Primary biliary cirrhosis (PBC) is characterized by an intense biliary inflammatory CD4(+) and CD8(+) T cell response. Very limited information on autoantigen-specific cytotoxic T lymphocyte (CTL) responses is available compared with autoreactive CD4(+) T cell responses. Using peripheral blood mononuclear cells (PBMCs) from PBC, we identified an HLA-A2-restricted CTL epitope of the E2 component of pyruvate dehydrogenase (PDC-E2), the immunodominant mitochondrial autoantigen. This peptide, amino acids 159-167 of PDC-E2, induces specific MHC class I-restricted CD8(+) CTL lines from 10/12 HLA-A2(+) PBC patients, but not controls, after in vitro stimulation with antigen-pulsed dendritic cells (DCs). PDC-E2-specific CTLs could also be generated by pulsing DCs with full-length recombinant PDC-E2 protein. Furthermore, using soluble PDC-E2 complexed with either PDC-E2-specific human monoclonal antibody or affinity-purified autoantibodies against PDC-E2, the generation of PDC-E2-specific CTLs, occurred at 100-fold and 10-fold less concentration, respectively, compared with soluble antigen alone. Collectively, these data demonstrate that autoantibody, helper, and CTL epitopes all contain a shared peptide sequence. The finding that autoantigen-immune complexes can not only cross-present but also that presentation of the autoantigen is of a higher relative efficiency, for the first time defines a unique role for autoantibodies in the pathogenesis of an autoimmune disease.

Show MeSH

Related in: MedlinePlus

PD5-specific CD8+ T cells from in vitro–cultured PBMCs. PBMCs from HLA-A2+ donors were cocultured with PD5-loaded APCs (autologous immature DCs) for 12 d, then restimulated with PD5 or a control peptide in the presence of brefeldin A, followed by intracellular staining for IFN-γ. (A) IFN-γ staining of samples from two PBC patients, PBC3 and PBC10, and two control donors, CL1 (alcoholic liver disease) and H1 (a healthy donor). Displayed in the dot plots are cells gated for lymphocyte population by forward-scattering and side-scattering and the CD4− population. The cells within the box are considered IFN-γ+. The number next to the box is the percentage of IFN-γ+ cells in the CD8+ T cell population. (B) Frequency of PD5-specific CD8+ T cells in PD5-stimulated PBMC cultures derived from all 20 donors. The cut off value for a positive response was determined as 0.150%, or 3 SD above the mean percentage of IFN-γ producing cells in all 20 samples restimulated with the control peptide. A significant number of IFN-γ–producing cells were detected after restimulation with PD5 in 10/12 (83%) PBC patients but in 0/8 control individuals (P < 0.0007).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2196012&req=5

fig1: PD5-specific CD8+ T cells from in vitro–cultured PBMCs. PBMCs from HLA-A2+ donors were cocultured with PD5-loaded APCs (autologous immature DCs) for 12 d, then restimulated with PD5 or a control peptide in the presence of brefeldin A, followed by intracellular staining for IFN-γ. (A) IFN-γ staining of samples from two PBC patients, PBC3 and PBC10, and two control donors, CL1 (alcoholic liver disease) and H1 (a healthy donor). Displayed in the dot plots are cells gated for lymphocyte population by forward-scattering and side-scattering and the CD4− population. The cells within the box are considered IFN-γ+. The number next to the box is the percentage of IFN-γ+ cells in the CD8+ T cell population. (B) Frequency of PD5-specific CD8+ T cells in PD5-stimulated PBMC cultures derived from all 20 donors. The cut off value for a positive response was determined as 0.150%, or 3 SD above the mean percentage of IFN-γ producing cells in all 20 samples restimulated with the control peptide. A significant number of IFN-γ–producing cells were detected after restimulation with PD5 in 10/12 (83%) PBC patients but in 0/8 control individuals (P < 0.0007).

Mentions: In an effort to examine the disease specificity of the CD8+ T cells against the identified PDC-E2 epitope (159–167) peptide, PBMCs from all 20 HLA-A2+ donors (12 PBC patients, PBC 1 to PBC 12; four patients with other chronic liver diseases CL1 to CL4, and four healthy donors H1 to H4) were cultured with autologous APCs loaded with the PD5 peptide. On day 12 the cells were restimulated with PD5 or a control peptide in the presence of brefeldin A, followed by intracellular IFN-γ staining. A significant number of IFN-γ–producing cells were detected after restimulation with PD5, but not with the control peptide, in 10 out of 12 (83%) PBC patients but none of the eight control individuals (P < 0.0007). Representative data from two PBC patients (PBC3 and PBC10) and two control individuals (CL1 and H1) are shown in Fig. 1 A, and the results of all 20 individuals tested are summarized in Fig. 1 B. The fact that PD5-specific CD8+ T cells can be expanded from the majority (10/12) of PBC patients but none of the control individuals strongly suggests that a CD8+ T cell response against the autoreactive epitope on PDC-E2 is associated with PBC.


Identification of HLA-A2-restricted CD8(+) cytotoxic T cell responses in primary biliary cirrhosis: T cell activation is augmented by immune complexes cross-presented by dendritic cells.

Kita H, Lian ZX, Van de Water J, He XS, Matsumura S, Kaplan M, Luketic V, Coppel RL, Ansari AA, Gershwin ME - J. Exp. Med. (2002)

PD5-specific CD8+ T cells from in vitro–cultured PBMCs. PBMCs from HLA-A2+ donors were cocultured with PD5-loaded APCs (autologous immature DCs) for 12 d, then restimulated with PD5 or a control peptide in the presence of brefeldin A, followed by intracellular staining for IFN-γ. (A) IFN-γ staining of samples from two PBC patients, PBC3 and PBC10, and two control donors, CL1 (alcoholic liver disease) and H1 (a healthy donor). Displayed in the dot plots are cells gated for lymphocyte population by forward-scattering and side-scattering and the CD4− population. The cells within the box are considered IFN-γ+. The number next to the box is the percentage of IFN-γ+ cells in the CD8+ T cell population. (B) Frequency of PD5-specific CD8+ T cells in PD5-stimulated PBMC cultures derived from all 20 donors. The cut off value for a positive response was determined as 0.150%, or 3 SD above the mean percentage of IFN-γ producing cells in all 20 samples restimulated with the control peptide. A significant number of IFN-γ–producing cells were detected after restimulation with PD5 in 10/12 (83%) PBC patients but in 0/8 control individuals (P < 0.0007).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196012&req=5

fig1: PD5-specific CD8+ T cells from in vitro–cultured PBMCs. PBMCs from HLA-A2+ donors were cocultured with PD5-loaded APCs (autologous immature DCs) for 12 d, then restimulated with PD5 or a control peptide in the presence of brefeldin A, followed by intracellular staining for IFN-γ. (A) IFN-γ staining of samples from two PBC patients, PBC3 and PBC10, and two control donors, CL1 (alcoholic liver disease) and H1 (a healthy donor). Displayed in the dot plots are cells gated for lymphocyte population by forward-scattering and side-scattering and the CD4− population. The cells within the box are considered IFN-γ+. The number next to the box is the percentage of IFN-γ+ cells in the CD8+ T cell population. (B) Frequency of PD5-specific CD8+ T cells in PD5-stimulated PBMC cultures derived from all 20 donors. The cut off value for a positive response was determined as 0.150%, or 3 SD above the mean percentage of IFN-γ producing cells in all 20 samples restimulated with the control peptide. A significant number of IFN-γ–producing cells were detected after restimulation with PD5 in 10/12 (83%) PBC patients but in 0/8 control individuals (P < 0.0007).
Mentions: In an effort to examine the disease specificity of the CD8+ T cells against the identified PDC-E2 epitope (159–167) peptide, PBMCs from all 20 HLA-A2+ donors (12 PBC patients, PBC 1 to PBC 12; four patients with other chronic liver diseases CL1 to CL4, and four healthy donors H1 to H4) were cultured with autologous APCs loaded with the PD5 peptide. On day 12 the cells were restimulated with PD5 or a control peptide in the presence of brefeldin A, followed by intracellular IFN-γ staining. A significant number of IFN-γ–producing cells were detected after restimulation with PD5, but not with the control peptide, in 10 out of 12 (83%) PBC patients but none of the eight control individuals (P < 0.0007). Representative data from two PBC patients (PBC3 and PBC10) and two control individuals (CL1 and H1) are shown in Fig. 1 A, and the results of all 20 individuals tested are summarized in Fig. 1 B. The fact that PD5-specific CD8+ T cells can be expanded from the majority (10/12) of PBC patients but none of the control individuals strongly suggests that a CD8+ T cell response against the autoreactive epitope on PDC-E2 is associated with PBC.

Bottom Line: Very limited information on autoantigen-specific cytotoxic T lymphocyte (CTL) responses is available compared with autoreactive CD4(+) T cell responses.Furthermore, using soluble PDC-E2 complexed with either PDC-E2-specific human monoclonal antibody or affinity-purified autoantibodies against PDC-E2, the generation of PDC-E2-specific CTLs, occurred at 100-fold and 10-fold less concentration, respectively, compared with soluble antigen alone.The finding that autoantigen-immune complexes can not only cross-present but also that presentation of the autoantigen is of a higher relative efficiency, for the first time defines a unique role for autoantibodies in the pathogenesis of an autoimmune disease.

View Article: PubMed Central - PubMed

Affiliation: Division of Rheumatology, Allergy and Clinical Immunology, University of California at Davis School of Medicine, Davis, CA 95616, USA.

ABSTRACT
Primary biliary cirrhosis (PBC) is characterized by an intense biliary inflammatory CD4(+) and CD8(+) T cell response. Very limited information on autoantigen-specific cytotoxic T lymphocyte (CTL) responses is available compared with autoreactive CD4(+) T cell responses. Using peripheral blood mononuclear cells (PBMCs) from PBC, we identified an HLA-A2-restricted CTL epitope of the E2 component of pyruvate dehydrogenase (PDC-E2), the immunodominant mitochondrial autoantigen. This peptide, amino acids 159-167 of PDC-E2, induces specific MHC class I-restricted CD8(+) CTL lines from 10/12 HLA-A2(+) PBC patients, but not controls, after in vitro stimulation with antigen-pulsed dendritic cells (DCs). PDC-E2-specific CTLs could also be generated by pulsing DCs with full-length recombinant PDC-E2 protein. Furthermore, using soluble PDC-E2 complexed with either PDC-E2-specific human monoclonal antibody or affinity-purified autoantibodies against PDC-E2, the generation of PDC-E2-specific CTLs, occurred at 100-fold and 10-fold less concentration, respectively, compared with soluble antigen alone. Collectively, these data demonstrate that autoantibody, helper, and CTL epitopes all contain a shared peptide sequence. The finding that autoantigen-immune complexes can not only cross-present but also that presentation of the autoantigen is of a higher relative efficiency, for the first time defines a unique role for autoantibodies in the pathogenesis of an autoimmune disease.

Show MeSH
Related in: MedlinePlus