H. Stein

Human dendritic reticulum cells of lymphoid follicles: their antigenic profile and their identification as multinucleated giant cells.

SummaryThe B-dependent areas of human lymphoid tissue contain non-lymphoid, non-phagocytic cells known as dendritic reticulum cells (DRC). These cells can be detected only very occasionally in routinely stained histologic sections. Recently we were able to overcome this limitation by preparing a monoclonal antibody, termed R 4/23, that reacts selectively with DRC. Thus by using an optimized immunoperoxidase method applied to frozen sections, it is possible to detect DRC in situ. To determine the antigenic profile of DRC, serial frozen sections of human tonsils were immunostained with R 4/23 and a large panel of other monoclonal antibodies or conventional antisera. In addition, touch imprints of tonsils and cytocentrifuge slides of cell suspensions with increased concentrations of DRC were immunostained with these reagents. DRC proved to be positive for μ, γ, α, κ and λ chains, complement component C3b, C3b receptors, C3d receptors, HLA-A,B,C antigens, human Ia-like antigens, common ALL antigen (cALLa), and antigens that are characteristic of the monocyte/macrophage lineages. DRC did not express δ chains, T cell antigens, or antigens that are expressed on interdigitating reticulum cells (IDC) and Langerhans cells. DRC in touch imprints and suspensions prepared from hyperplastic tonsils were found to be giant cells often with 10 or more nuclei. In certain cases of follicular hyperplasia and of centroblastic-centrocytic lymphoma, DRC with several nuclei were also detectable in situ.These results show that (1) the phenotype of DRC differs from that of all other cell types in lymphoid tissue, (2) this phenotype most nearly resembles that of cells of the monocyte/macrophage series, thus suggesting that DRC are related to these cell lineages, and (3) DRC are multinucleated giant cells.

MALIGNANT LYMPHOMAS OF B-CELL TYPE

Abstract The immunoglobulin (1g) concentration in tissue homogenates of 28 malignant lymphomas and 3 myelomas was examined and compared with that of 25 normal lymph-node homogenates. 15 cases showed normal or reduced Ig values and 16 showed a significant increase of one of the Ig types. IgA had increased in 1, IgG in 4, and IgM in 11 cases. Histologically these tumours were classified as: reticulum-cell sarcoma (4), chronic lymphocytic leukaemia (3), multiple myeloma (3), lymphoblastic lymphosarcoma (2), lymphocytic lymphosarcoma (1), lymphatic plasma-cell leukaemia (1), unclassifiable lymphoma (2).

An immunohistological study of the cellular constituents of Hodgkin's disease using a monoclonal antibody panel

Cryostat sections of lymphoid tissue from 44 cases of Hodgkins disease were analysed by immunoperoxidase staining using a panel of monoclonal antibodies which included reagents reactive with T cells and their subsets, B cells, HLA‐DR, Ig, dendritic reticulum cells and C3b receptor. A wide spectrum of immunohistological patterns was observed ranging from cases in which T cells were numerous (B cells being absent or present in only small numbers) to cases in which very prominent B cell follicles were present. These follicles contained a meshwork of dendritic reticulum cells and were composed of polyclonal B cells (as assessed by light chain expression). T cells were present in small numbers within these B cell follicles, often clustered in a thin rim around individual Reed‐Sternberg and Hodgkins cells. All B cell‐rich cases were examples of lymphocyte predominant Hodgkins disease. Assessment of the T cell helper/suppressor ratios was hindered by the fact that both anti‐helper antibodies (OKT4 and anti‐Leu 3a) reacted with macrophages. However the majority of cases appeared to contain a normal excess of T helper cells. HLA‐DR was strongly expressed in T cell rich areas, on Reed‐Sternberg and Hodgkins cells, on vascular endothelium and on numerous infiltrating cells in the fibrous tissue areas in cases of nodular sclerosing disease. Reed‐Sternberg and Hodgkins cells were not labelled by either anti‐fibronectin or by antibodies reactive with dendritic reticulum cells (anti‐C3b receptor and antibody R4/23).

Demonstration of lysozyme, α1-antichymotrypsin, α1-antitrypsin, albumin, and transferrin with the immunoperoxidase method in lymph node cells

SummaryThe immunoperoxidase method was used to investigate the presence of intracytoplasmic lysozyme, α1-antichymotrypsin (α1-ACT), α1-antitrypsin (α1-AT), transferrin, and albumin in hyperplastic and inflamed human lymph nodes. Lysozyme was demonstrated in eosinophils, neutrophils, histiocytes, in epithelioid cells, mast cells, and some lining cells of lymph node sinuses. α1-ACT was detectable in many, but not all histiocytes that stained for lysozyme, and in sinus histiocytes, epithelioid cells, and mast cells, but not in neutrophils or eosinophils. α1-AT was demonstrable in mast cells, neutrophils, and some epithelioid cells, but not in histiocytes. Transferrin was found in mast cells, but not in any of the other cell types investigated. Albumin was detectable in a few epithelioid cells and giant cells of the Langhans type. Lysozyme, α1ACT, α1-AT, transferrin, and albumin were never demonstrable in interdigitating reticulum cells, dendritic reticulum cells, or lymphoid cells.

Immunohistological analysis of human lymphoid tissue by double immunoenzymatic labelling.

SummaryThe increasing number of antigens detectable in human lymphoid tissue (particularly since the advent of monoclonal antibodies) makes it necessary to have techniques available for studying the relative distribution patterns of pairs of antigens in tissue sections. Double immunoenzymatic labelling (using peroxidase and alkaline phosphatase) offers a number of advantages over double immunofluorescence, including the fact that the two antigens can be visualised simultaneously (rather than sequentially) and that the labels are permanent.In studying paraffin-embedded human lymphoid tissue an important application of the double immunoenzymatic technique lies in distinguishing Ig-positive cells containing exogenous Ig (which causes mixed κ/λ staining) from cells containing endogenous Ig (which stain for only a single light chain class). In addition double staining of paraffin sections for IgG and IgM has been used to show that “switch” cells containing both these classes of heavy chain are rare in reactive lymphoid tissue.The potential scope of the double immunoenzymatic technique has been extended by showing that the procedure is applicable to cryostat sections (in which antigenic reactivity is better preserved than in paraffin sections) and by adapting it for use with monoclonal antibodies (by preparing “monoclonal PAP” complexes).

Cytogenetic findings in T-zone lymphoma

SummaryFive cases of T-zone lymphoma were investigated with histologic, immunologic, and cytogenetic methods. The chromosome analyses were performed on lymphoma cells prepared immediately after removal of the lymph nodes. The chromosomes involved in structural rearrangements were nos. 1, 2, 3, 4, 14, and Y. Numbers 3, 5, 6, and 13 were lost by some tumors, and nos. 3 and 9 were gained. Chromosome 3 was involved most often in structural and numerical aberrations, whereas 14q+ markers occurred in only one case. The importance of multidisciplinary studies is pointed out.

Lysosomal acid phosphatase: Activity and isoenzymes in separated normal human blood cells

The present study was devised to investigate the activity and isoenzymes of lysosomal acid phosphatase in individual normal human blood cells, including the T- and B-population of lymphocytes, with the aim to contribute to the classification of haematopoietic neoplasias on the basis of cell specific isoenzyme patterns. Platelets, erythrocytes, granulocytes, monocytes and T-lymphocytes were isolated from blood by gradient centrifugation or immune adsorption. B-lymphocytes were obtained from human tonsils. After purity control and isolation of lysosomes the concentration of acid phosphatase was assayed using the conventional spectrophotometric method. Isoenzymes were separated by isoelectric focusing on polyacrylamide thin layer slabs. Monocytes revealed the highest activity with 14 mU/10(7) cells, about three times more than granulocytes. T-lymphocytes showed an activity of 2.85 mU/10(7) cells and B-lymphocytes of 1.83 mU/10(7) CELLS. The lowest activity was found in platelets with 0.08 mU/10(7) cells. Granulocytes showed 12 isoenzyme bands, whilst the number for monocyte, B-lymphocytes, T-lymphocytes and platelets were respectively 11, 12, 1 and 4 isoenzyme bands. Thus it became evident that the different blood cell populations can be distinguished on the basis of their acid phosphatase isoenzyme pattern.

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.

Makroglobulinbildende chronische lymphatische Leukämie ohne Makroglobulinämie

SummaryA lymph node of a patient suffering from chronic lymphocytic leukemia was investigated using histological, cytochemical and electron microscopic techniques. The remaining lymph node tissue and the blood were also examined for their immunoglobulin content. The immunoglobulin analysis revealed an excessive increase of IgM in the lymph node, while there was a reduced serum IgM level. A morphological equivalent of the increased IgM was constituted by the deposit of PAS-positive cytoplasm inclusions, which were embedded in distended cisternae of the ergastoplasm of the neoplastic lymph node cells. “Secretory” lymphatic cells containing ergastoplasm without inclusion bodies otherwise dominated the electron microscopic picture.The existence of a B-cell lymphoma producing IgM of the secretory type is thus demonstrated. The lack of emission of IgM into the blood has to be interpreted as the result of a defect in the secretory mechanism. As the IgM was found in the form of 8 S-IgM in the tissue, it can be assumed that a disturbance of the polymerisation of the IgM-subunits into the 19 S-pentamer existed, the result of which was the absence of secretion of IgM into the blood.ZusammenfassungBei einem Fall von chronischer lymphatischer Leukämie wurden histologische, cytochemische und elektronenmikroskopische Untersuchungen des Lymphknotens sowie Immunglobulinbestimmungen des Lymphknotenhomogenates und des Blutes durchgeführt.Die Immunglobulinbestimmungen deckten eine hochgradige IgM-Vermehrung im Lymphknoten bei IgM-Verminderung im Blut auf. Ein morphologisches Äquivalent dieser IgM-Vermehrung stellte die Ablagerung PAS-positiver Cytoplasmaeinschlüsse in den neoplastischen Lymphknotenzellen dar. Diese Einschlüsse waren in Ergastoplasmasäcken dieser Zellen lokalisiert. Im übrigen beherrschten ergastoplasmahaltige „sekretorische“ lymphatische Zellen ohne cytoplasmatische Einschlüsse das elektronenmikroskopische Bild.Es handelte sich also um ein IgM-bildendes B-Zellenlymphom vom sekretorischen Typ. Die unterbliebene Abgabe der Makroglobuline in das Blut ist als Sekretverhaltung zu deuten. Da das IgM als 8 S-IgM im Gewebe vorlag, nehmen wir an, daß hier eine Polymerisationsstörung der IgM-Untereinheiten zu dem 19 S-Pentamer bestand, als deren Folge die unterbliebene Sekretion von IgM ins Blut anzusehen ist.

An immunohistological study of reactive lymphoid tissue

The aim of this study was to document the patterns of cytoplasmic Ig heavy and light chain expression in reactive lymphoid tissue, using single and double immunoenzymatic labelling techniques. This investigation was undertaken, firstly, to provide information on whether the normal counterparts of high grade lymphoma cells (e.g. centroblasts, immunoblasts) ever express more than one light or heavy chain (as has been noted in the past for lymphomas) and also, secondly, to seek evidence of intraclonal ‘switching’ from cytoplasmic IgM to cytoplasmic IgG expression. Paraffin embedded sections, all showing substantial reactive changes, were analysed by means of immunoperoxidase stains for the three major immunoglobulin classes (IgG, IgM and IgA), both light chain classes and J chain. In addition, double immunoenzymatic labelling techniques were used to search for cells showing simultaneous expression of kappa and lambda light chains and cells expressing mu and gamma heavy chain. Large transformed lymphocytes showing cytoplasmic Ig‐staining in the pulp and interfollicular areas often have nuclear morphology indistinguishable from germinal centre centroblasts. There was no evidence of primitive appearing IgM‐positive cells and IgG‐positive cells of more mature morphology. In addition, immunoenzymatic staining showed that cells simultaneously expressing both IgG and IgM are only rarely encountered. When such cells were detected, the morphology was not that of a blast cell, but rather of a plasma cell containing Russell bodies. Hence it is suggested that cytoplasmic IgM switching to IgG is rarely detected by immunohistological methods in reactive tissue. Double staining for kappa and lambda revealed that cells simultaneously expressing both light chain types were not detected even among cells showing the most primitive morphology.