Monoclonal antibodies reveal multiple forms of expression of human microsomal epoxide hydrolase.
In a previous study, we developed five kinds of monoclonal antibodies against different portions of human mEH: three, anti-N-terminal; one, anti-C-terminal; one, anti-conformational epitope. Using them, we stained the intact and the permeabilized human cells of various kinds and performed flow cytometric analysis. Primary hepatocytes and peripheral blood mononuclear cells (PBMC) showed remarkable differences. On the surface, hepatocytes exhibited 4 out of 5 epitopes whereas PBMC did not show any of the epitopes. mEH was detected inside both cell types, but the most prominent expression was observed for the conformational epitope in the hepatocytes and the two N-terminal epitopes in PBMC. These differences were also observed between hepatocyte-derived cell lines and mononuclear cell-derived cell lines. In addition, among each group, there were several differences which may be related to the cultivation, the degree of differentiation, or the original cell subsets. We also noted that two glioblastoma cell lines reveal marked expression of the conformational epitope on the surface which seemed to correlate with the brain tumor-associated antigen reported elsewhere. Several cell lines also underwent selective permeabilization before flow cytometric analysis, and we noticed that the topological orientation of mEH on the ER membrane in those cells was in accordance with the previous report. However, the orientation on the cell surface was inconsistent with the report and had a great variation between the cells. These findings show the multiple mode of expression of mEH which may be possibly related to the multiple roles that mEH plays in different cells.
Department of Microbiology, Faculty of Medicine, Saitama Medical University, Moroyama-cho, Iruma-gun, Saitama 350-0495, Japan.
Hongying
H
Duan
Akira
A
Takagi
Hidekazu
H
Kayano
Isamu
I
Koyama
Christophe
C
Morisseau
Bruce D
BD
Hammock
Toshitaka
T
Akatsuka
D000328:Adult; D000368:Aged; D000911:Antibodies, Monoclonal / Q000276:immunology*; D002460:Cell Line; D045744:Cell Line, Tumor; D000939:Epitopes; D004851:Epoxide Hydrolases / Q000235:genetics / Q000276:immunology*; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D015971:Gene Expression Regulation, Enzymologic; D005909:Glioblastoma / Q000201:enzymology / Q000276:immunology; D022781:Hepatocytes / Q000201:enzymology / Q000276:immunology; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000201:enzymology / Q000276:immunology; D008297:Male; D008875:Middle Aged
D000911:Antibodies, Monoclonal; D000939:Epitopes; D004851:Epoxide Hydrolases