H. J. Ree

Lectin histochemistry of malignant tumors. I. Peanut agglutinin (PNA) receptors in follicular lymphoma and follicular hyperplasia: an immunohistochemical study.

Peanut agglutinin (PNA) receptors were studied in 37 cases of reactive follicular hyperplasia and 66 follicular lymphomas, using the unlabeled peroxidase–antiperoxidase (PAP) method on paraffin embedded material. Based on the binding sites of the lectin, positively stained cells were easily recognized as either cytoplasmic receptor‐positive (CR+) or surface receptor‐positive (SR+) cells. In the lymph node specimens, CR+ cells corresponded to macrophage–histiocytes and possibly dendritic reticulum cells; SR+ cells corresponded to lymphoid cells. Three categories of CR+ cells were noted: large, medium, and small. The large CR+ cells were present in most germinal centers from reactive nodes, but were virtually absent in neoplastic follicles. Varying numbers of medium and small CR+ cells were seen in reactive as well as neoplastic follicles. SR+ cells were present in both follicular lymphoma (64%) and follicular hyperplasia (19%). In neoplastic follicles, SR+ cells were distributed uniformly throughout every follicle in the node revealing no relation to the orientation of the node. In reactive follicles, however, the occurrence of SR+ cells was not only infrequent, but also focal, and was often associated with the polarity of the follicles. The uniform distribution of SR+ tumor cells produced a characteristic staining pattern of neoplastic follicles which, along with the disappearance of the large CR+ cells, would provide an additional feature useful in the differential diagnosis of neoplastic from reactive follicles.

Macrophage-histiocytes in Hodgkin's disease the relation of peanut-agglutinin-binding macrophage-histiocytes to clinicopathologic presentation and course of disease

The authors studied the occurrence of peanut agglutinin (PNA)‐binding cells in paraffin‐embedded specimens of 145 patients with Hodgkins disease (HD). The staining reaction of lymphocytes was consistently negative. A positive staining reaction was observed in two types of cells: macrophagehistiocytes (M‐H), and Reed‐Sternberg (R‐S) cells and their variants. Diffuse or globular cytoplasmic staining was found in M‐H, which was easily distinguished from a unique “cell surface and cytoplasmic” staining pattern of R‐S and related cells. Thus defined, M‐H were numerous in lymphocyte depletion and mixed cellularity, less common in lymphocyte predominance, and least frequent in the nodular sclerosis type. Numerous M‐H correlated with B‐symptoms and a poor response to therapy. Among the asymptomatic patients with localized disease at presentation, the presence of large numbers of M‐H was associated with a high incidence of relapse within 2 years of therapy. These findings suggest that the number of non‐neoplastic M‐H in HD may be an important determinant in the clinical presentation and course of disease. Peanut agglutinin staining may be useful for the detection of M‐H in routine diagnosis and classification of HD, which has not been feasible by conventional methods.

Lectin histochemistry of malignant tumors II. Concanavalin A: A new histochemical marker for macrophage–histiocytes in follicular lymphoma

Concanavalin agglutinin (Con A) binding sites were studied in paraffin embedded lymph node specimens of reactive follicular hyperplasia (12 cases) and follicular lymphoma (37) using the avidin‐biotin‐peroxidase complex method, and the results were compred with those of Peanut agglutinin (PNA) and lysozyme stains. Very similar to the PNA stain, two categories of Con A receptor sites were observed: cytoplasmic and cell surface. In the reactive lymph nodes, the cells showing cytoplasmic receptor sites (CR+ cells) corresponded to macrophage–histiocytes and possibly dendritic reticulum cells in the H & E stained sections, while those showing cell surface receptor sites (SR+) corresponded to lymphoid cells. Unlike the PNA binding, however, the staining reaction of SR+ lymphoid cells was weak, and another staining pattern, a dot‐like stain, was observed in some lymphocytes, both SR+ and SR–. In follicular lymphomas, CR+ histiocytes were distinctly displayed within the follicular centers in 25 of 37 cases, including 12 cases in which PNA stains on adjacent or nearby sections were negative for intrafollicular macrophage–histiocytes. Similarly, Con A stains were positive for the intrafollicular CR+ cells in four of the five cases in which lysozyme stains were negative. Many of these intrafollicular CR+ cells contained inclusion‐like cytoplasmic globules and/or vacuoles, a hallmark of the large CR+ cells of germinal centers. These observations suggest that macrophage–histiocytes of presumably germinal center origin are retained in neoplastic follicular centers in varying degrees, and Con A might be a useful marker for macrophage–histiocytes in paraffin‐embedded routine pathological specimens, in addition to the currently accepted markers, PNA and lysozyme.

Lectin distinction of benign from malignant histiocytes

Ricinus communis agglutinin (RCA) staining of malignant histiocytosis (two cases) and tissue infiltrates of monoblastic leukemia revealed two distinct macrophage‐histiocyte types: one with cytoplasmic staining (RCA+), the other without (RCA‐). RCA+ cells corresponded to benign‐appearing histiocytes without nuclear atypia, whereas RCA‐ cells were cytologically malignant. In one case of malignant histiocytosis, many RCA‐ tumor cells were surrounded by a thin cytoplasmic extension of RCA+ cells, which suggested an interaction between the tumor cells and stromal macrophage‐histiocytes. These observations support the view that most benign‐appearing histiocytes in malignant histiocytosis are reactive in nature and suggest that RCA staining can be useful in distinguishing between benign and malignant disorders of macrophage‐histiocytes. Cancer 56: 2046‐2050, 1985.

Sclerosis in diffuse histiocytic lymphoma: A clinicopathologic study of 25 cases

Sclerosis was observed in the lymph node specimens of 26 of 57 (46%) patients with diffuse histiocytic lymphoma. Two major types of sclerosis were observed: “compartmentalizing” (56%), in which the tumor was divided into many small compartments by anastomosing, hyalinized stroma; and “diffuse” (44%), in which the sclerosis occurred without evident partitioning of the tumor. The hyalinized stroma of the compartmentalizing sclerosis was frequently continuous with small vessels accompanied by aggregates of small lymphocytes, suggesting that the sclerosis was related to the occurrence of small lymphocyte‐associated postcapillary venules. Compartmentalizing sclerosis was further divided into two groups: The first group (even pattern) was characterized by an orderly occurrence of the stromal network with no evidence of distortion, evenly spaced tumor cells showing no axis in their arrangement, and a sharp demarcation of tumor cells from the stroma. The second group (uneven pattern) was marked by either a disorderly occurrence of the stromal network with distortion of the overall pattern, tumor cells arranged along an axis, or poor demarcation of tumor cells from the stroma with individual envelopment of tumor cells by the stroma. Approximately two‐thirds of the patients (9/14) with the compartmentalizing sclerosis survived two years or more after diagnosis; most patients (10/11) with diffuse sclerosis died within two years. Compartmentalizing sclerosis, even pattern, was associated with a consistently favorable prognosis; the uneven pattern was not. This study indicates a marked variation in the survival of patients with diffuse histiocytic lymphoma with sclerosis and demonstrates that prognostic subgroups may be delineated by additional morphologic features.

Occurrence and patterns of muramidase containing cells in Hodgkin's disease, non-Hodgkin's lymphomas, and reactive hyperplasia

The occurrence and pattern of cytoplasmic muramidase containing histiocytes were studied by the unlabeled antibody peroxidase-antiperoxidase method in biopsy material from patients with Hodgkins disease, non-Hodgkins lymphomas, and reactive hyperplasia. The majority of lymph nodes from patients with Hodgkins disease, nodular lymphoma, and reactive hyperplasia gave positive staining reactions when tested in this manner. Differences in the staining pattern were observed for the different conditions studied. In general, stain positive cells occurred in one of the following four patterns: nodular, dispersed, aggregating without background stain, or aggregating with background stain (mottling pattern). The nodular and aggregating without background stain patterns were not specific and were seen in various conditions. The dispersed pattern, however, was observed only in some cases of non-Hodgkins diffuse lymphomas, suggesting a subgroup of tumors characterized by active participation of reactive histiocytes. The mottling pattern was virtually limited to Hodgkins disease. Since the mottling pattern appeared to be produced by virtue of a large amount of extracellular muramidase, the elevation of the serum muramidase level in Hodgkins disease may be related to enzymatically active secretory histiocytes. Moreover, the mottling staining pattern was observed frequently in the lymphocytic predominance and nodular sclerosis type of Hodgkins disease, but relatively infrequently in the mixed cellularity or lymphocytic depletion types, suggesting that the variation in histiocytic activity may be related to the course of the disease. The decreased staining reaction observed in the latter two categories could not be accounted for by a decrease in the numbers of histiocytic cells in hematoxylin and eosin stained sections, suggesting that release or synthesis may be defective in those unfavorable types of Hodgkins disease.

Anti-interleukin-1 reactive cells in Hodgkin's disease.

Constitutional symptoms (or B‐symptoms) of Hodgkins disease may be mediated by interleukin 1 (IL‐1), a product of macrophage‐histiocytes. To further study this relation, the authors examined the cells that reacted with anti‐human IL‐1 antibody in biopsy specimens from 140 untreated patients with Hodgkins disease (72 asymptomatic patients and 68 with B‐symptoms). Fever was the most common symptom, present in 57 of the 68 patients. Anti‐IL‐1 reactive cells were observed in 62 cases. A positive staining reaction was observed in three types of cells: Reed‐Sternberg and related (R‐S) cells (33 cases); small to medium cells of undetermined origin (18); and granulocytes (11). The staining was negative in 78 cases, including 42 with B‐symptoms. The majority of tumors (27/33) with positively stained R‐S cells were from asymptomatic patients. Most tumors (14/18) with positively stained small to medium sized cells were from patients with B‐symptoms. Large numbers of granulocytes were positively stained in five asymptomatic patients and six with B‐symptoms. The immunohistochemical demonstration of IL‐1‐bearing cells in tumors does not correlate with the manifestation of constitutional symptoms in Hodgkins disease.

Macrophage‐histiocyte lysozyme activity in relation to the clinical presentation of Hodgkin's disease: An immunohistochemical study

The clinical presentation of 71 untreated patients with Hodgkins disease was studied in relation to immunohistochemically demonstrable lysozyme in the lymph node biopsy material. Sixty‐one patients (86%) showed a positive staining reaction of varying degree, while ten (14%) showed no demonstrable lysozyme. The clinical features of lysozyme‐positive patients differed markedly from those of lysozyme‐negative patients. Stain‐positive patients were younger (29 vs. 46), were more often in clinical Stage I or II disease (69% vs. 10%, P< 0.001), and less frequently had constitutional symptoms (34% vs. 70%, P< 0.02). Moreover, within the stain‐positive group, patients who had the most intense staining reaction (mottling pattern) also had the most favorable clinical and histopathologic features at the time of diagnosis. The observations suggest that in Hodgkins disease the lysozyme secretory activity of macrophage‐histiocytes may be an important element of host resistance to neoplasia and that a depression of this secretory activity corresponds with disseminated disease.

Concanavalin A‐binding histiocytes in Hodgkin's disease: A predictor of early relapse

Staining with Concanavalin agglutinin (Con A) reveals a far greater number of macrophage‐histiocytes (M‐H) in paraffin sections than any other staining method. With Con A staining, the shapes of stromal M‐H are clearly visualized, thus enabling a study of their morphologic variations. Con A staining patterns were also unchanged in specimens left at room temperature for 24 to 28 hours before fixation. The appearance of Con A‐binding histiocytes was studied in tumors, recurrent as well as original, of 18 patients with biopsy‐proven early relapse (within 26 months of diagnosis), and compared with those of 26 patients who were in complete remission (lasting 48 months at the minimum). The early‐relapse patients were diagnosed from 1977 through 1984, and all received intensive combination chemotherapy. The relapse‐free patients were treated in various manners, and included six patients diagnosed in the 1960s who were treated with radiation alone. Three forms of Con A‐binding histiocytes were easily recognized: medium‐sized cells similar to those seen in reactive follicles, characterized by uniform nuclei and distinct, abundant cytoplasm (Type A); cells of varying size and shape with altered cytoplasm, rarefied and ragged with indistinct cell borders, or globular (Type B); and large cells, stellate or spindling (Type C). Large numbers of Type A cells were present in all tumors of the relapse‐free patients but were virtually absent in the original and recurrent tumors of the early‐relapse group. Conversely, Type B cells were rare in the relapse‐free group, but were the most common type in the patients with early relapse. Type C cells were not seen in the former group, but were present in the latter. These observations suggest that the morphologic variations of Con A‐binding histiocytes in Hodgkins disease are associated with tumor behavior. Con A staining, which can best depict stromal histiocytes in paraffin sections, may be used to identify patients at a high risk of early relapse. Cancer 58:87–95, 1986.

Macrophage-histiocytes in malignant lymphoma, small lymphocytic type (well-differentiated lymphocytic lymphoma).

The authors studied the occurrence of Ricinus communis agglutinin (RCA)‐binding macrophage—histiocytes in paraffin—embedded tumor tissue of 38 patients with malignant lymphoma, small lymphocytic type, a tumor of low‐grade malignancy. Thirty‐one patients (82%) had an indolent clinical course and were free of disease for a minimum follow‐up period of 24 months. However, seven patients (18%) died within 24 months of biopsy, and six of the seven patients died of rapid progression of their tumor despite intensive treatment. Histologically, the tumors of these six short‐term survivors were indistinguishable from those of the long‐term survivors. RCA staining of paraffin‐embedded tumor tissue of the 38 cases revealed three groups of tumors: (1) tumors with numerous (>10/high‐power field [HPF]) stromal macrophage—histiocytes (4 patients); (2) tumors with a moderate number (4–9/HPF) of macrophage—histiocytes (5 patients); (3) tumors with rare or no (0–3/HPF) macrophage—histiocytes, or only thin, anuclear variants (29 patients). Each of the six short‐term survivors had readily demonstrable RCA‐binding macrophage—histiocytes in their tumor; these were numerous in four and moderate in two. In contrast, macrophage—histiocytes were either rare or absent, or were anuclear variants, in 29 of the 31 patients who had an indolent clinical course. These observations suggest that in small lymphocytic type malignant lymphoma there is a subgroup characterized by an increased number of stromal macrophage—histiocytes and aggressive behavior of the tumor. Tumors of this subgroup can be detected by RCA staining.