High molecular weight vimentin complex is formed after proteolytic digestion of vimentin by caspase-3: detection by sera of patients with interstitial pneumonia.
In a previous study, we demonstrated anti-vimentin antibodies in sera of patients with interstitial pneumonia. We hypothesized that antibodies in sera might detect vimentin fragments formed during the process of apoptosis. To prove this, recombinant human vimentin was digested by recombinant human caspase-3 or caspase-8. Then, Western blotting using several commercially available antibodies against human vimentin or patients' sera which had anti-vimentin autoantibodies, was performed. As a result, after recombinant human vimentin was digested by caspase-3 or caspase-8, several vimentin fragments were formed and detected by 2 kinds of monoclonal anti-vimentin antibodies (clone 3B4 and clone V9) as well as by polyclonal sheep anti-human vimentin antibody. It was demonstrated that high molecular weight vimentin was formed after the digestion of vimentin by caspase-3, which was only detected by patients' sera. The high molecular weight vimentin was not formed after digestion of vimentin by caspase-8. Our present results show that high molecular weight vimentin was formed after the digestion of vimentin by caspase-3. In addition, it is suggested that this high molecular weight vimentin acted as an autoantigen to form anti-vimentin autoantibody in vivo.
Fujita
Jiro
J
First Department of Internal Medicine, Kagawa Medical University, Kagawa 761-0793, Japan. jiro@kms.ac.jp
Bandoh
Shuji
S
Yang
Yu
Y
Wu
Fei
F
Ohtsuki
Yuji
Y
Yoshinouchi
Takeo
T
Ishida
Toshihiko
T
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D017209:Apoptosis; D001323:Autoantibodies / Q000097:blood / Q000276:immunology*; D001324:Autoantigens / Q000096:biosynthesis / Q000737:chemistry* / Q000276:immunology; D001327:Autoimmune Diseases / Q000097:blood / Q000276:immunology*; D015153:Blotting, Western; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D017563:Lung Diseases, Interstitial / Q000097:blood / Q000276:immunology*; D046911:Macromolecular Substances; D008297:Male; D008875:Middle Aged; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000378:metabolism; D012756:Sheep; D014746:Vimentin / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D001324:Autoantigens; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D014746:Vimentin; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases