Cloning of variable regions of an antibody that reacts with the soluble fraction of human liver cells and its possible value in chronic liver disease
✍ Scribed by H Saito; S Tada; H Ebinuma; K Atsukawa; T Masuda; Y Inagaki; K Tsuchimoto; T Morizane; H Ishii
- Publisher
- John Wiley and Sons
- Year
- 1996
- Tongue
- English
- Weight
- 940 KB
- Volume
- 23
- Category
- Article
- ISSN
- 0270-9139
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✦ Synopsis
other mouse immunoglobulins. These findings suggest A gene encoding the variable regions of the heavy and that a human antibody with the same idiotype as a light chains of a mouse monoclonal antibody designated mouse monoclonal antibody that reacts with human H2, which specifically reacts with human liver cells, was liver cells, can be detected in patients with chronic liver cloned into a phagemid vector. The clone of the variable disease, suggesting that autoimmunity may be partly reregion was designed to be expressed as a separate prosponsible for these diseases. (HEPATOLOGY 1996;23:1498tein, the structure of which is the same as that of the 1506.) mouse antibody. The cloned phage protein specifically reacted with anti-idiotypic antibodies produced in rabbits against the original mouse antibody, and this reac-Autoimmune responses against liver cells or their tion was specifically blocked by the original antibody. components have been implicated in some forms of
The soluble protein, expressed as a fusion protein, was chronic hepatitis. Since Mackay et al. 1 described an detected as a single 30-kd band on sodium dodecyl sulautoimmune hepatitis with associated antibodies fate polyacrylamide gel electrophoresis (SDS-PAGE) and specifically bound to an anti-H2 idiotypic antibody against organ-and species-nonspecific nuclear antias determined by Western blot analysis. Sera of patients gens, various autoantibodies have been detected in the with various diseases were assayed for antibodies to sera of patients with autoimmune hepatitis. These auanti-H2 by sandwich enzyme-linked immunosorbent toantibodies can be valuable diagnostic markers. These assay (ELISA). Only sera from patients with chronic autoantibodies also have been detected in some paliver disease reacted strongly. This binding was specifitients with chronic hepatitis (CH) caused by hepatitis cally blocked by the cloned soluble protein. The nucleoviruses. Although there have been criticisms of the imtide sequences of the variable regions were determined munologic findings in patients with viral or autoimby the dideoxy chain-termination method, and the semune CH, 2 the existence of autoantibodies in the sera quences were approximately 95% identical to those of of CH patients suggests that an autoimmune mechanism may be responsible for the CH in these patients.
We have previously developed a murine monoclonal Abbreviations: CH, chronic hepatitis; MAb, monoclonal antibody; LSP, antibody (MAb), designated H2, 3 that was produced by liver-specific membrane lipoprotein; ASG-R, anti-asialoglycoprotein-receptor fusing P3-NS1 cells with lymph node cells from BALB/ antibody; anti-H2ID, anti-H2-idiotype; Ig, immunoglobulin; PCR, polymerase chain reaction; VH, heavy chain variable gene; VL, light chain variable gene; c mice sensitized with a fractionated soluble phase of ScFv, single-chain fragment containing VH and VL regions; rf, recombinant a human liver homogenate. H2-MAb reacts specifically phage; PBS, phosphate-buffered saline; RT, room temperature; ELISA, enwith normal human liver cells in immunohistochemiszyme-linked immunosorbent assay; HRP, horseradish peroxidase; E-tag, the try studies. 3,4 If an autoimmune mechanism is involved peptide tag; SDS, sodium dodecyl sulfate; PAGE, polyacrylamide gel electroin patients with chronic liver disease, we postulate that phoresis; AIH, autoimmune hepatitis; HCC, hepatocellular carcinoma; LSIA, liver-specific idiotype-bearing antibody; ANA, antinuclear antibody; SMA;
an antibody against the liver might be present in these anti-smooth muscle antibody; LKM-1; liver-kidney microsome-1 antibody; nt, patients. Antibodies against liver-specific membrane not tested.