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The presence of pan-cadherin and the MEP-1 antigen in normal and diseased rat lung was established by employing immunoperoxidase and double label fluorescence techniques. The binding of a mouse monoclonal antibody (MEP-1) reacting specifically with type 1 pneumocytes was assessed on paraffin sections of normal specimens as well as those with pulmonary fibrosis induced by bleomycin or radiation treatment. In injured alveolar epithelium, a diminished type 1 cell and a foca1 type 11 cell immunoreactivity was found. Electron microscopy of immunogold-labelled lung tissue confirmed the type 1 cell specificity of MEP-l. In severely injured pulmonary parenchyma MEP-Inegative areas occurred, which were also negative with the type 11 pneumocyte marker Maclura pomifera lectin. Similarly, a polyclonal pan-cadherin antibody uniformly decorated luminal surfaces of alveoli except the type 11 pneumocytes. Furthermore, pleural mesothelial cells, bronchiolar epithelial cells, endothelial cells of large blood vessels and alveolar macrophage surfaces exhibited pan-cadherin immunoreactivity. After injury, a remarkable loss of pan-cadherin immunoreactivity in the MEP-1-positive type 1 epithelial cells was detectable. These findings suggest that characterization of normal alveolar epithelial cells and monitoring of the epithelial remodelling in pulmonary pathohistology are sufficiently described by the antibodies MEP-1 anti pancadherin.

To elucidate the deposition of advanced glycation end products (AGEs) in aortic atherosclerosis, aortic walls were obtained from 25 autopsy cases and examined immunohistochemically and immunoelectron microscopically with a monoclonal antibody specific for AGEs, 6D12. Among the autopsy cases, atherosclerotic lesions were found in the aortas of 22 cases and were composed of diffuse intimal thickening, fatty streaks, atherosclerotic plaques, and/or complicated lesions. In these cases, intracellular AGE accumulation was demonstrated in the intimal lesions of aortic atherosclerosis in 12 cases. Compared with the diffuse intimal thickening, intracellular AGE accumulation was marked in the fatty streaks and atherosclerotic plaques. Immunohistochemical double staining with 6D12 and monoclonal antibodies for macrophages or muscle actin or a polyclonal antibody for scavenger receptors demonstrated that the AGE accumulation in macrophages or their related foam cells was marked in the diffuse intimal thickening and fatty streak lesions and that almost all macrophages and macrophage-derived foam cells possessed scavenger receptors. Immunoelectron microscopic observation revealed the localization of 6D12-positive reaction in lysosomal lipid vacuoles or electron-dense granules of the foam cells. These results indicate that AGE accumulation occurs in macrophages, smooth muscle cells, and their related foam cells.

Malignant fibrous histiocytoma (MFH) was induced in rats by injection of 9,10-dimethyl-1,2-benzanthracene. Using cell suspensions prepared from the heterotransplanted nude mouse tumor as immunogen, a monoclonal antibody, (MAb), MEP-1, against fibroblastlike MFH tumor cells was generated. In the primary rat tumors and transplanted rat or nude mouse tumors, MEP-1 reacted specifically with the fibroblastlike cells but not with the histiocytelike cells or xanthoma cells. Anti-rat macrophage MAbs RM-1 and TRPM-3 did not stain the fibroblastlike cells, but both were reactive with the histiocytelike cells. Double stainings with both MEP-1 and RM-1 or TRPM-3 did not detect any double positive cells. Immunoelectron microscopy using these MAbs showed that the fibroblastlike cells were the major cell component of the primary and transplanted rat tumors and that their cell membrane was stained positively with MEP-1, but not for RM-1 or TRPM-3. By the double staining method using a MAb against prolyl 4-hydroxylase beta and MEP-1 or TRPM-3, this enzyme was demonstrated in MEP-1-positive cells but not in TRPM-3-positive cells. Results obtained by DNA cytofluorometry with 4,6-diamidino-2-phenylindole dihydrochloride staining or by the combined method of DNA cytofluorometry and indirect immunofluorescence, using MEP-1, RM-1, and TRPM-3, indicate that MEP-1-positive cells are neoplastic cells of rat MFH having proliferation activity. In the transplanted nude mouse tumors, no differentiation of MEP-1-positive rat tumor cells into histiocytelike cells was detected, and all histiocytelike cells were immunostained by F4/80 and most of them were positive for M5/114. These results suggest that fibroblastlike cells and intermediate cells are tumor cells of 9,10-dimethyl-1,2-benzanthracene-induced rat MFH showing differentiation toward fibroblasts and that histiocytelike cells are infiltrated macrophages.

An anti-macrophage monoclonal antibody (mAb), RbM2, was produced using thioglycolate-elicited rabbit peritoneal macrophages as immunogen. The immunohistochemical approach revealed an intense reactivity of this mAb to macrophages, particularly those engaged in phagocytosis, in different organs and tissues, and to peripheral blood monocytes. Immunoelectron microscopy demonstrated reaction products of RbM2 in the lysosomes of macrophages and monocytes. This selective reactivity with the mAb was further confirmed in various experiments by endocytosis of Latex particles with different diameters or IgG-coated sheep erythrocytes. The results indicate that RbM2 recognizes a lysosomal membrane antigen of 50,000 molecular weight (MW). In contrast, dendritic cells, such as follicular dendritic cells (FDC) of lymphoid follicles, interdigitating cells (IDC) of lymphoid T zones, or epidermal Langerhans' cells, were not reactive with the antibody. Thus, RbM2 is useful not only in differentiating the phagocytic macrophages from the immunologically accessory dendritic cell populations but also in identifying lysosomes and their related structures in macrophages.

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