Heinz J. Radzun

Ki-M2R, a new specific monoclonal antibody, discriminates tissue macrophages from reticulum cells and monocytes in vivo and in vitro.

Utilizing rat peritoneal macrophages as the immunogen, a new monoclonal antibody enabling differential monitoring of the mononuclear phagocyte system (MPS) by immunohistochemistry has been raised. Designated Ki‐M2R, this antigen could be detected with the immune alkaline phosphatase reaction on all macrophages including those of bone marrow, lymphatic sinuses, lymphoid follicles, splenic red pulp, and von Kupffer cells of the liver, as well as on macrophages of connective tissue, renal interstitial tissue, serous cavities, and gastrointestinal tract. Langerhans cells—the MPS‐derived reticulum cells of the epidermis—interdigitating reticulum cells, and dendritic reticulum cells of lymphoid follicles were invariably negative. Blood monocytes were rendered positive only after evolving into macrophages upon appropriate stimulation. Thus, Ki‐M2R selectively labels monocytes after transformation into macrophages.

Kinetics of Kupffer cells as shown by parabiosis and combined autoradiographic/immunohistochemical analysis

SummaryLeucocytes from syngeneic rats were labeled with tritiated thymidine and donor and recipient rats were connected by a bilateral arteriovenous shunt. Based on the time-dependent label index and labeling intensity, it was concluded that Kupffer cells, the resident macrophages of the liver, have a half-life of 12.4 days and originate from monocytes undergoing one mitosis within 8.4 days after immigration into the liver. The labeled cells were easily identified as Kupffer cells by their selective immunoreactivity with the monoclonal antibody Ki-M2R which is specific for phagocytosing macrophages in the rat. The applicability of combined autoradiography/immunohistochemistry for the identification of other poorly defined macrophage subpopulations is shown.

Activity and isoenzymes of acid phosphatase in human B-cell lymphomas of low-grade malignancy: a novel aid in the classification of malignant lymphoma.

Activity and isoelectric focusing (IEF) pattern of lysosomal acid phosphatase (E.C.3.1.3.2.) were investigated in 55 cases of low‐grade malignant B‐cell lymphoma, classified as chronic B‐lymphocytic leukemia (B‐CLL), centroblastic/centrocytic follicular lymphoma (CB/CC), lymphoplasmacytic/lymphoplasmacytoid lymphoma (Immunocytoma, IC), and plasmacytoma (PC), applying the criteria of the Kiel classification. The results show (1) that the four lymphoma types present a characteristic range of enzyme activity in an increasing order: B‐CLL, CB/CC, IC, and PC. B lymphocytes, germinal center cells, and plasmacytes are the main constituents of these lymphomas. This sequence might reflect one possible mode of B‐cell transformation into plasmacytes traversing an amplification stage in germinal centers under normal conditions. (2) All cases showed the basic IEF pattern of normal B lymphocytes with 12 bands localized in three regions between pH 6.1 and 3.9. This finding supports the B‐cell origin and the close phenotypical relationship among the investigated lymphomas. (3) The IEF patterns of B‐CLL and CB/CC did not differ from that of normal B lymphocytes, whereas two additional isoenzymes were encountered in cases of IC and seven in PC; this suggests that the higher enzyme activity of IC and PC is at least partly due to the appearance of “new” isoenzymes. The results support the validity of the underlying classification and indicate the individuality, B‐cell origin, and close relationship among the four lymphoma entities investigated.

Modulation of c‐fms Proto‐Oncogene Expression in Human Blood Monocytes and Macrophages

The gene product of the c‐fms proto‐oncogene is a transmembrane protein with tyrosine‐kinase activity that is obviously related to the receptor for the colony‐stimulating‐factor CSF‐1. By Northern blot analysis, we investigated the expression of the cellular counterpart of v‐fms in purified normal human blood mononuclear cells and different macrophage populations. The proto‐oncogene c‐fms expression was demonstrable in blood monocytes but not in blood lymphocytes. Short‐term cultivated blood monocytes exhibited an increased expression of c‐fms in comparison to freshly isolated blood monocytes, possibly due to a temporary down regulation of c‐fms during the separation procedure of blood monocytes. A comparably high rate of fms‐RNA expression was found in most of the analyzed samples of resident peritoneal macrophages, while resident alveolar macrophages showed a considerably lower level of c‐fms expression. In this, alveolar macrophages resembled long‐term cultivated adherent blood monocytes, which showed a down regulation of c‐fms expression. By correlating these data obtained by Northern blot analysis with phenotypic properties of the analyzed monocyte/macrophage populations, it is concluded that different levels of c‐fms expression in monocytes/macrophages correspond to their stage of differentiation and maturity.

Analysis of acid esterase activity and polymorphism as an aid for the classification of human low‐grade malignant Non‐Hodgkin's lymphomas

Investigating a link between enzyme histochemical and recent immunohistochemical results, the authors studied the activity and the polymorphism of acid esterase (EC 3.1.1.6) in well defined human B‐cell lymphomas. Twelve cases of chronic B‐lymphocytic leukemia, 18 cases of centroblastic/centrocytic follicular lymphoma, and 17 cases of lymphoplasmacytic/lymphoplasmacytoid lymphoma, as diagnosed according to Kiel classification, were subjected to enzyme assay and isoelectric focusing of acid esterase. Enzyme values revealed no characteristic distribution among the lymphoma entities. The isoenzyme pattern, specific to normal human B‐lymphocytes, were regularly detectable in all lymphoma entities. The results document the B‐cell origin of the analyzed subsets of B‐cell malignancies, although distinctive acid esterase patterns were lacking. The prevalence of three anodal isoenzymes in cases of chronic B‐lymphocytic leukemia, hardly detectable in other lymphoma entities, were interpreted as the expression of a clonal proliferation, arrested at a certain differentiation stage of B‐cells common for the majority of the tumor cells. Hence, further evidence is provided supporting the view that the studied entities represent B‐cell neoplasias at different stages of differentiation expressing a variety of different markers.

Human macrophage hybrid forming spontaneous giant cells

SummaryThymidine kinase-deficient clones of the human monocyte/ macrophage cell line U-937 were established and used for fusion experiments with separated normal human blood monocytes. A hybrid (H 29) was generated during HAT-selection procedure, about 50% of which formed spontaneous giant cells, as shown by morphological, immunocytochemical, and chromosomal analyses. It is concluded that giant cells originate from monocytes and display mitotic activity. Macrophage hybrids are the basic requirement for the elucidation of monocyte/macrophage heterogeneity and immortalization of their functional properties.

Enzyme polymorphism in the classification of human malignant lymphoma

SummaryRecent studies on the polymorphism of lysosomal hydrolases have shown that all individual blood cell types in the human being possess their own isoenzyme pattern. In the present study acid phosphatase activity of normal human B-lymphocytes and of four different types of low-grade malignant non-Hodkings lymphomas according to the Kiel classification was estimated. In addition, the isoenzyme pattern of AcP was investigated by isoelectric focusing. Chronic lymphocytic leukemia of B-cell type (N=9) and centroblastic centrocytic follicular lymphoma (N=10) demonstrated significantly lower values than lymphoplasmacytic/lymphoplasmacytoid lymphomas (N=28) and plasmacytomas (N=8). The isoenzyme pattern of normal human B-lymphocytes comprised 12 bands between pH 6.3 and 3.85. This basical pattern was shared by all four lymphoma entities. Only lymphoplasmatic/lymphoplasmacytoid lymphoma and plasmacytoma revealed additional bands, which probably account for the higher net enzyme activity in these cases.

Six new monoclonal antibodies to serous, mucinous, and poorly differentiated ovarian adenocarcinomas.

Six monoclonal antibodies directed against ovarian adenocarcinoma were generated by use of 100,000 X g supernatants of Triton‐X‐100 solubilized extracts of ovarian serous adenocarcinoma as the antigen source. Immunoperoxidase preparation of frozen‐sections and routinely processed paraffin section specimens revealed a highly restricted reactivity of these antibodies when tested with adult (n = 2) and fetal (n = 3) tissue types. Coreactivities were occasionally observed with epithelia of the kidney, mammary gland, and pancreas. One monoclonal antibody, Ki‐OC I‐6‐2, cross‐reacted only with epididymal epithelia. No coreaction occurred with normal tissue of the ovary, Fallopian tube, or uterus. All antibodies were additionally tested on 74 cases of nonovarian malignancies, 15 cases of ovarian metastases of nonovarian carcinomas, and 114 specimens of ovarian neoplasms other than carcinomas. Ki‐OC I‐6‐2 had no cross‐reactivity with these tumors except for one case of renal cell carcinoma. This monoclonal antibody recognized serous, mucinous, and poorly differentiated adenocarcinoma cell types. None of the six antibodies reacted with clear cell or endometrioid carcinoma. All were found to be of the IgG‐1 subclass. The tumor antigen to which Ki‐OC I‐6‐2 immunoreacted was estimated to have a molecular weight of 80 kilodaltons (KD).

Immunophenotyping of acute non-lymphoblastic leukaemias.

SummaryUsing monoclonal antibodies specific for myelomonocytic cells, 40 non-lymphoblastic leukaemias were analysed applying immunostaining to cytospin preparations. Based on the reactivity patterns six groups of acute non-lymphoblastic leukaemias could be determined, mirroring the bimodal differentiation pathway of myelomonocytic cells. Comparative enzyme cytochemical analysis did not render a clear cut correlation and discrimination of the immunocytochemically defined groups. It is concluded that only the application of a broad panel of immunocytochemical and enzyme cytochemical methods allow a sound subdivision and diagnosis of acute non-lymphoblastic leukaemias.