sIg Expression Research Articles

Immunoglobulin somatic variation; studies of receptor editing in a human B cell lymphoma.

In order to study mechanisms of immunoglobulin somatic variation in committed immunoglobulin (Ig) expressing B cells, we used the fluorescent activated cell sorter to isolate rare variants from a surface Ig positive (sIg+) diffuse large cell B lymphoma cell line (mu lambda+). These variants were either negative for sIg expression (sIg-) or expressed sIg which differed from the original parental tumor cell line, both in idiotypes and Ig lambda isotypes (sIg+Id-). In the following report we review the results from the studies of these variants. DNA analysis showed that all variants had new Ig lambda gene rearrangements, which had occurred either on a previously excluded allele, or on the productively rearranged allele of the parental cell line. The sIg- variants, which had undergone nonfunctional Ig lambda rearrangements on the expressed parental allele, and thereby deleted the productive rearrangement, continued to functionally rearrange the same allele and regenerated sIg expression. While the parental cell line expressed low levels of the recombination activating genes, RAG1 and RAG2, expression of these genes were considerably upregulated in both the immunoglobulin negative and the idiotypic variants. Somatic immunoglobulin V gene hypermutation did not contribute to the observed immunoglobulin somatic variation. These results demonstrate that, through differential expression of the RAG genes, sIg+ B cells are able to somatically alter their sIg receptors through secondary Ig lambda gene rearrangements. This mechanism may allow B cells with non-selectable, or auto-reactive, antigen receptors to alter these receptors (receptor editing). Ongoing Ig gene rearrangement may limit the usefulness of immunotherapeutic approaches directed at the antigen receptor in some diffuse large cell lymphomas.

New Cell Line from Hairy-Cell Leukaemia Producing lnterleukin-6 after Epstein-Barr Virus Immortalization

A hairy-cell leukaemia (HCL) cell line, HCL-O, was established from the peripheral blood of a 62-year-old Japanese patient with a unique variant of HCL strongly expressing CD21, the receptor for the Epstein-Barr virus (EBV). The HCL-O cells expressed antigens similar and dissimilar to those expressed with the original hairy cells. The HCL-O cells were more mature than the original cells in their degree of B-cell differentiation, as indicated by a decrease of CD19 and surface immunoglobulin (sIg) expression together with the appearance of CD38 and cytoplasmic Ig (cIg). In addition, the cells expressed CD11c recognized by Leu-M5, a monoclonal antibody usually positive for HCL. Their karyotype and Ig gene rearrangement pattern were identical to those of the original cells. The EBV genome was detected in the HCL-O cells but not in the original cells. The HCL-O cells spontaneously produced a large quantity of interleukin-6 (IL-6) in the conditioned medium, whereas IL-6 serum level was not so high. These findings indicate that the HCL-O cell line is derived from the leukaemic hairy cells and possibly, in vitro EBV infection took place easily in the original hairy cells through their CD21, resulting in subsequent immortalization. IL-6 production by HCL-O cells may be induced or enhanced by EBV, and the secreted IL-6 might play a role in their own growth or differentiation.

Accessory molecules that influence signaling through B lymphocyte antigen receptors.

The B lymphocyte antigen receptor complex, commonly called the B cell receptor (BCR) complex, consists of surface immunoglobulin (sIg) and heterodimers of the Igα (CD79a, mb1) and Igβ (CD79b, B29) phosphoglycoproteins. Recent reviews detail pertinent findings on the BCR complex.1 The structure of the BCR complex and some cell-surface molecules that influence signaling via the BCR are shown in Figure 1. The clonotypic Ig receptor has only a very short cytoplasmic tail and therefore must rely on the invariant members of the BCR complex to transmit signals to the cytosol after receptor crosslinking. Igα/Igβ associate with sIgM and sIgD and are both necessary and sufficient for the expression of sIg. This heterodimer is analogous to the TCR CD3ε/δ or CD3ε/γ heterodimers,2 which, like Igα/Igβ, (1) contain subunits with a single extracellular Ig-like domain; (2) are phosphorylated on tyrosine (CD3ε and CD3ζ) after crosslinking of antigen receptors; and (3) contain within their cytoplasmic tail a single antigen receptor homology 1 (ARH1) motif, D/E-X7-D/E-X2-Y-X2-L-X7-Y-X2-L/I (X = any amino acid). Regions within the transmembrane domain of sIgM are required for the release of [Ca2+]i or internalization of bound antigen.3 Mutations within the transmembrane domain of sIgM that inhibit the activation of new protein tyrosine phosphorylation (PTP) and release of [Ca2+]i also uncouple sIgM from Igα/Igβ.4 However, even though such a mutant sIgM does not associate with Igα/Igβ, when crosslinked it still induces some new PTP.4 Both Sanchez et al.4 and Kim et al.5 reported that surface chimeric fusion proteins expressing the cytoplasmic tails of Igα vs. Igβ differ in their ability to transmit signals: the Igα but not the Igβ tail can induce new PTP, results consistent with studies suggesting that Igα and not Igβ strongly associates with the protein tyrosine kinase (PTK) p53/56Lyn (Lyn).6 Matsuuchi et al.7 found that sIgM expression could be reconstituted in a pituitary cell line with Igα/Igβ coexpression, but that Igα/Igβ were not sufficient to reconstitute a complete signal through sIgM. Thus, IgM interaction with Igα/Igβ is critical for signaling but other factors may also be required.

Intracellular Induction Pathways for CD23 Antigen and Surface Immunoglobulins in Human Tonsillar B Cells: The Roles of Protein Kinase C and Tyrosine Kinaxe-Mediated Signals

The expression of CD23 on human tonsillar B cells is increased following treatment with interleukin 4 (IL-4) or 12-O-tetradecanoylphorbol 13-acetate (TPA), while that of surface immunoglobulins (sIgs) is increased by IL-4 but decreased by TPA. This suggests that the signaling by these effectors may result from distinct second messenger-generating systems. In this study, we attempted to elucidate the signal transduction pathways responsible for the expression of CD23 and sIgs by using different protein kinase C (PKC) and tyrosine kinase (TK) inhibitors. Our results showed that B cells expressed varying amounts of sIgs depending on different activators and inhibitors. Sphingosine, a PKC inhibitor, almost completely reversed the TPA-induced decrease in sIgM and sIgD expression. Other PKC inhibitors, e.g., H7 and staurosporine, had similar but less profound effects. In comparison, the up-regulation of CD23 by IL-4 and TPA was only partially blocked by these PKC inhibitors. TK inhibitors, such as herbimycin A and genistein, decreased both the IL-4- and TPA-induced CD23 expression by 50-80%, but had modest effects on sIgs expression. These findings indicate that CD23 and sIgs expression is regulated by independent pathways: PKC is important for the regulation of sIgs expression while the signals through TK pathways might play the major role in CD23 expression.

Modulation of B Lymphocyte Differentiation by Calcitonin Gene-Related Peptide (CGRP)

The presence of calcitonin gene-related peptide (CGRP) in nerve endings in lymphoid organs, around blood vessels, and in bone marrow suggests that it might influence the function and differentiation of lymphoid cells. Previous studies identified specific CGRP receptors on mature T and B lymphocytes and on 70Z/3 pre-B cells. In these studies, it was found that CGRP stimulated a rapid, prolonged elevation of cAMP with in 70Z/3 cells with an ED50 of approximately 20 fM. Following CGRP treatment, cAMP levels peaked within 5 min and were still elevated after 60 min. The effect of CGRP on surface immunoglobulin (sIg) expression was examined by treating 70Z/3 cells with CGRP, or combinations of CGRP and LPS, and then measuring slg expression by FACS. When 70Z/3 cells were treated with LPS, CGRP, or calcitonin for 48 hr, only LPS induced the expression of sIg, increasing the percentage of cells expressing sIg from less than 10% positive in untreated cells to 80-98% positive. Subsequent experiments examined the effect of CGRP on LPS-induced sIg. CGRP inhibited LPS-induced sIg expression at concentrations ranging from 10-15 to 10-7M. The maximal inhibition was observed at CGRP concentrations ranging from 10-10 to 10-8M, and the maximal reduction of sIg expression was about 40%. The inhibitory effect of CGRP was specific in that it could not be mimicked by calcitonin and could be blocked by the CGRP receptor antagonist CGRP8-37. A similar dose-dependent inhibitory effect on LPS induction was observed in 70Z/3 cells treated with LPS and dibutyryl cAMP, suggesting that the inhibitory effect of CGRP on sIg expression is mediated by stimulation of intracellular cAMP. The inhibitory effect of CGRP on LPS induction of sIg appears to be mediated by a reduction in the expression of both μ and κ mRNA. When μ and κ expression were examined by Northern blot analysis, it was found that CGRP caused a 50% reduction in the amount of μ and κ mRNA induced by LPS. The ability of CGRP to inhibit differentiation of 70Z/3 cells and sIg expression provides evidence that CGRP can influence the differentiation of lymphopoeitic precursors.

Up-regulated recombination-activating gene expression in sIg- variants of a human mature B cell line undergoing secondary Ig lambda rearrangements in cell culture.

To study the role of surface immunoglobulin (sIg) expression in the control of rearrangement activity at the Ig light chain loci, we established rare sIg- clones (lambda-) from a human sIg+ B cell line (mu lambda+). Upon expansion of these sIg- clones, surface immunofluorescence analysis revealed a gradual emergence of sIg+ subpopulations, differing from the original tumor cell line both in their idiotypes and C lambda isotypes. DNA analysis revealed that this sIg heterogeneity resulted from a process of ongoing Ig lambda rearrangements. That is, one of the Ig lambda rearrangements in the parental cell line was replaced by novel Ig lambda rearrangements in the sIg- clones, which in turn were replaced by yet additional Ig lambda rearrangements in the sIg+ variants. Northern analysis demonstrated that while the expression of the recombination-activating genes RAG1 and RAG2 was relatively low in the parental cell line, their expression was significantly increased in both the sIg- variants and their sIg+ progenies. We thus describe a human mature B cell line, in which differential RAG expression allows sIg heterogeneity to be generated through secondary Ig lambda gene rearrangements. Our results indicate that the induction of RAG expression may be inversely associated with sIg expression, but that sIg expression, alone, is not sufficient to down-regulate this expression.

Tyrosine phosphorylation of CD19 in pre-B and mature B cells.

Cross-linking of B cell surface immunoglobulins (sIg) results in activation of mature B cells and stimulates a molecular signaling mechanism for antigen-specific B cell expansion and differentiation. This signaling pathway is dependent on tyrosine (Tyr) phosphorylation and results in the activation of sIg-associated src family kinases and p72SYK. Rapid Tyr phosphorylation occurs on multiple protein substrates. Here we show that activation of B cells by cross-linking sIg results in an increase in Tyr phosphorylation of the lineage-restricted B cell surface antigen CD19, and show that it is a major substrate of activated Tyr kinase following sIg stimulation. Lower levels of constitutive CD19 Tyr phosphorylation occurred in most sIg+ mature B cell lines examined and in normal dense tonsillar B cells. We also find that when CD19 is Tyr-phosphorylated it becomes competent to interact with SH2 domains suggesting a mechanism whereby, following B cell activation, CD19 could be linked to intracellular signaling pathways. In sIg- pre-B cell lines, CD19 was expressed but was not constitutively phosphorylated on tyrosine. Upon CD19 cross-linking, Tyr phosphorylation of CD19 was induced in sIg- pre-B cell lines. CD19 cross-linking also directly induced Tyr phosphorylation of CD19 and other substrates in mature B cells. The ability of CD19 to signal in the absence of sIg expression may provide important stimulation in pre-B cell development.

The effect of IL-4 on the phenotype of a human B-cell lymphoma line (Farage) lacking immunoglobulin expression

Farage cells do not express surface immunoglobulins (sIg) but display a high level of CD19, CD21, CD22, CD23, CD39, CD40 B-cell antigens, and various adhesion proteins, such as CD11a (LFA-1), CD29 (VLA-4), CD44, CD54 (ICAM-1), and CD58 (LFA-3). The phenotype of Farage resembled that of EBV-LCL but differed from the phenotype of Burkitt's lymphoma lines, which were CD39 − CD44, expressed a high level of CD38, and either lacked CD21 or were weakly positive. Exposure to IL-4 augmented the concentrations of CD23 and of adhesion proteins on the surface of Farage cells but diminished the expression of CD21, CD22, and CD38. IL-4 did not induce the expression of sIg on Farage cells and failed to affect the level of HLA-DR. IL-2 and TPA did not alter the level of CD21 and adhesion proteins on Farage cells. Although IL-4 induced unique changes of the antigenic pattern in Farage, no significant effect on the phenotype of Burkitt's lymphoma lines was detected after IL-4 treatment. The present study indicates that the responsiveness of B cells to IL-4 is not determined by the expression of sIg but rather is associated with the antigenic profile characteristic for non-germinal center B lymphocytes.

Mouse rosettes and surface immunoglobulin in small lymphocytic lymphoma. Importance in immunophenotyping and differential diagnosis

Cell suspensions from lymphoid tissue of 82 small lymphocytic lymphoma (SLL), 8 intermediate lymphocytic lymphoma (ILL), 286 other B-non-Hodgkin's lymphoma (B-NHL), and 248 reactive lymphadenopathy (RLA) cases were analyzed to evaluate the diagnostic significance of mouse-rosette (M-rosette) assay, and surface immunoglobulin clonality (SIg) and level of expression. In SLL, 55 were M-rosette positive (67.07%) and 72 SIg positive (87.8%), with weak fluorescence in 63 and strong fluorescence in 9 cases. Of 10 SIg-negative cases, 9 were M-rosette positive; of 27 M-rosette-negative cases, 26 were SIg positive. Seven of the nine cases with strong fluorescence were M-rosette positive. In other B-NHL, 252 were M-rosette negative (88.11%) and 245 SIg positive (85.66%), with strong fluorescence in 211 and weak fluorescence in 34 cases. Thirty-two of the 34 cases with weak fluorescence were M-rosette negative. Of the RLA cases, 213 were M-rosette negative (85.89%) and 1 SIg positive (0.4%). The study demonstrated the independent expression of M-rosettes and SIg in SLL and their complementary role in diagnosis. It showed that positive results for M-rosettes and weak fluorescence are characteristic of SLL, that M-rosette negativity and strong fluorescence are characteristic of other B-NHL, and that M-rosette negativity and polyclonal SIg are characteristic of RLA. In 26 cases with paired data for CD5, M-rosettes, and SIg, a positive result for M-rosettes was superior to CD5 in differentiating SLL from other B-NHL. Intermediate lymphocytic lymphoma frequently showed weak SIg fluorescence and M-rosette negativity.

Expression of 5'-nucleotidase (CD73) related to other differentiation antigens in leukemias of B-cell lineage

Ecto-5'nucleotidase (5'NT; CD73) expression was studied with a monoclonal antibody (7G2) and a radiochemical assay and compared with the expression of other antigens in B-cell-lineage leukemias on cells from 100 leukemic patients and two cell lines. A B-cell origin was confirmed by the expression of CD19 and HLA-DR. Four stages of B-cell leukemias were defined: stage I (pro-B) as CD10-, cytoplasmic mu- (c mu-), surface Ig- (sIg-); stage II (cALL) as CD10+/c mu-/sIg-; stage III (pre-B) as CD10+ or -/c mu+/sIg-; and stage IV (B) as CD10-/c mu-/sIg+. A linear correlation was found between immunohistochemical and radiochemical determination of 5'NT (r = .86). 5'NT expression was low in T-cell leukemias and stage I, high in stages II and III, and low again in stage IV of B-cell leukemias. 5'NT expression was not related to c mu, CD20, CD21, CD22, CD34, and terminal deoxynucleotidyl transferase (TdT) expression, but was significantly related to CD10 and inversely related to kappa/lambda expression. However, the 5'NT activity in CD10+ leukemias (stages II and III) shows a very wide range. Within the group of CD10+ leukemias no differences were detected between 5'NT+ and 5'NT- cells in their expression of other B-cell antigens. We conclude that the place of 5'NT in leukemias corresponding to early stages of B-cell development has been characterized. 5'NT is expressed in CD10+ stages and decreases before the expression of sIgs. Future studies should make clear whether a high expression of this enzyme in CD10+ stages is a normal maturation phenomenon or a malignant phenomenon.

Expression of 5' -Nucleotidase (CD73) Related to Other Differentiation Antigens in Leukemias of B-Cell Lineage

Abstract Ecto-5′nucleotidase (5′NT; CD73) expression was studied with a monoclonal antibody (7G2) and a radiochemical assay and compared with the expression of other antigens in B-cell-lineage leukemias on cells from 100 leukemic patients and two cell lines. A B-cell origin was confirmed by the expression of CD19 and HLA-DR. Four stages of B-cell leukemias were defined: stage I (pro-B) as CD10-, cytoplasmic mu- (c mu- ), surface Ig- (sIg-); stage II (cALL) as CD10+/c mu-/sIg-; stage III (pre-B) as CD10+ or -/c mu+/sIg-; and stage IV (B) as CD10-/c mu-/sIg+. A linear correlation was found between immunohistochemical and radiochemical determination of 5′NT (r = .86). 5′NT expression was low in T-cell leukemias and stage I, high in stages II and III, and low again in stage IV of B-cell leukemias. 5′NT expression was not related to c mu, CD20, CD21, CD22, CD34, and terminal deoxynucleotidyl transferase (TdT) expression, but was significantly related to CD10 and inversely related to kappa/lambda expression. However, the 5′NT activity in CD10+ leukemias (stages II and III) shows a very wide range. Within the group of CD10+ leukemias no differences were detected between 5′NT+ and 5′NT- cells in their expression of other B-cell antigens. We conclude that the place of 5′NT in leukemias corresponding to early stages of B-cell development has been characterized. 5′NT is expressed in CD10+ stages and decreases before the expression of sIgs. Future studies should make clear whether a high expression of this enzyme in CD10+ stages is a normal maturation phenomenon or a malignant phenomenon.

Concomitant delineation of surface Ig, B‐cell differentiation antigens, and HLADR on lymphoid proliferations using three‐color immunocytometry

Accurate and consistent enumeration of B-cell subpopulations in lymphoid tissue was achieved through multiparameter three-color immunofluorescence and flow cytometric analysis (FCM). Phycoerythrin (PE)-anti-CD19 (Leu12) and biotinylated anti-HLADr/streptavidin-Duochrome (PE/Texas Red), used in conjunction with polyclonal fluorescein isothiocyanate (FITC) conjugated anti-surface immunoglobulin (SIg) antibodies, effectively separated non-specific binding and background fluorescence from true B-cell surface FITC immunofluorescence, while concomitantly analyzing for HLADr and CD19 phenotypic expression/deletion. Autofluorescence was measured to establish a fluorescence threshold. A second control measured non-specific binding of isotypic control mouse Ig and non-immune rabbit IgG. Cell suspensions from 128 samples of various lymphoid proliferations were studied. In 116 of the 128 samples, kappa/lambda ratios determined by flow cytometry correlated well with immunocytology results obtained using cytospins from the same cell suspension and with histopathologically established diagnosis. Clonality and lineage as defined immunotypically by flow cytometry was concordant with genotypic results in 64 of the 67 cases evaluated. SIg, HLADr, and CD19 deletions were demonstrated by flow cytometry in 8, 4, and 1 case(s), respectively. Discordance was usually attributable to selective loss of large neoplastic cells in flow cytometry specimens or absent expression of SIg by some cytoplasmic Ig (CIg+) lymphomas.

Radiation-induced surface IgG modulation: protein kinase C involvement.

Although radiation-induced loss of surface IgG (s-IgG) expression on murine B cells is known to be dependent on intact energy metabolism and integrity of the cytoskeleton, the exact mechanism of this radiation effect is not known. Evidence reported here shows that inhibition of protein kinase C (PKC) by H-7 impairs the radiation-induced s-IgG modulation, whereas addition of HA-1004, which preferently inhibits c-AMP-dependent protein kinase, shows only minor effects. On the other hand PMA, a PKC activator, mimics the radiation effect, and H-7 but not HA-1004 inhibits the PMA-induced loss of s-IgG expression. Therefore it is suggested that PKC is involved in the modulation of s-IgG induced by irradiation on B cells. The possibility of membrane participation in this event is discussed.

Up-regulation and down-regulation of cell surface and mRNA expression of CD5 antigen by various humoral factors on murine 70Z/3 pre-B cell leukemia cell line: IL-4 down-regulates CD5 antigen expression

CD5, a pan-T cell antigen, is expressed on a minor subset of normal B lymphocytes and on cells of most B lineage tumors or transformed B cells in both man and animal models. In the present study, the effects of various humoral factors on CD5 expression by cells of a subcloned 70Z/3 murine pre-B leukemia cell line were investigated. Among the humoral factors studied, only LPS up-regulated CD5 expression on 70Z/3 cells (three- to fourfold) in a dose-dependent manner. However, this up-regulatory effect of LPS was not observed when cells were cultured in serum-free medium. NZB-serum factor (NZB-SF), a cytokine we have identified and shown to enhance the maturation and proliferation of immature B cells, synergistically enhanced CD5 expression in the presence of suboptimal doses of LPS. IL-4 down-regulated CD5 expression by 70Z/3 cells induced by LPS or LPS plus NZB-SF in a dose-dependent manner. IL-4 also suppressed spontaneous CD5 expression by 70Z/3 cells. No other cytokine tested showed an inhibitory effect. LPS, IFN-γ, NZB-SF, and IL-1 enhanced sIg expression on 70Z/3 cells and their action on sIg expression was not inhibited by IL-4. Thus, the down-regulatory action of IL-4 on CD5 expression appeared specific for this antigen. IFN-γ, which inhibits IL-4 induced CD23 and DR expression on B cells, does not abolish the down-regulatory action of IL-4 on CD5 expression by 70Z/3 cells. Changes in mRNA levels on coding CD5 were also examined following the incubation of 70Z/3 cells (24 hr) in the presence of humoral factors which can influence CD5 Ag expression. The levels of mRNA for CD5 Ag were moderately increased in the presence of LPS and NZB-SF. IL-4 appeared to suppress the actions of NZB-SF and LPS at least in part by reducing the levels of mRNA encoding CD5.

Correlation of clinical outcome with the degree of surface immunoglobulin (sIg) expression in B-cell chronic lymphocytic leukemia.

Peripheral blood from 121 consecutive adult patients with B-cell chronic lymphocytic leukemia (B-CLL) was immunophenotyped, and three groups with differing surface immunoglobulin (sIg) expression patterns were identified: Group A, 82 patients (68%) with weak sIg expression in greater than 20% of the circulating lymphocytes; Group B, 16 patients (13%) with either undetectable sIg or weak sIg expression limited to less than 20% of the lymphocytes; and Group C, 23 patients (19%) with strong sIg expression in greater than 20% of the lymphocytes. The histories of these patients were reviewed retrospectively, and the median follow-up in Groups A, B, and C was nine, ten, and nine years, respectively. Disease progression, defined as the development of cytopenia(s) (hemoglobin less than or equal to 100 g/L [less than or equal to 10 g/dL] or platelet count less than 100 X 10(9)/L [less than 100 X 10(3) microL]) and/or the development of newly palpable splenomegaly or lymphadenopathy, was least in Group B (6%) versus Group A (34%) versus Group C (30%) (P = 0.08). The incidence of hypogammaglobulinemia among patients who had protein electrophoresis performed was 7% in Group B versus 46% in Group A versus 45% in Group C (P = 0.036). The authors conclude that adult patients with B-CLL in whom sIg is undetectable or weakly expressed in less than 20% of the circulating lymphocytes may have an even more indolent clinical course than those with more widespread sIg expression.

Antigen-independent regulation of cytoplasmic calcium in B cells with a 12-kDa B-cell growth factor and anti-CD19.

Increases in cytoplasmic free calcium ([Ca2+]i) can be induced in resting B cells either by a low molecular weight (12-kDa) B-cell growth factor (LMW-BCGF) or by crosslinking the B-cell antigen CD19 with monoclonal antibody (mAb). LMW-BCGF causes a slow [Ca2+]i increase in peripheral blood and tonsillar B cells but has no effect on [Ca2+]i in resting T cells. B-cell [Ca2+]i responses mediated by anti-surface immunoglobulin (sIg) or anti-CD19 are potentiated by LMW-BCGF, but anti-sIg and anti-CD19 do not show additive [Ca2+]i responses. LMW-BCGF- and anti-CD19-induced [Ca2+]i signals are similar to the sIgM or sIgD-mediated signals in that they are inhibited by prior treatment with phorbol 12-myristate 13-acetate. However, LMW-BCGF- and CD19-mediated signals do not depend on the expression of sIg, since they were also observed on sIg-B-cell precursor acute lymphoblastic leukemia (ALL) cells. Both anti-CD19 and LMW-BCGF stimulated in vitro colony formation by ALL cells and showed additive effects when used together. [Ca2+]i responses to LMW-BCGF or CD19 cross-linking were also evident on certain pre-B-cell and lymphoma B-cell lines.

A unique antigen on mature B cells defined by a monoclonal antibody.

A novel 42,000 dalton antigen (MB-1) expressed by mature human B cells in blood and tonsil was identified and characterized by utilizing a hybridoma monoclonal antibody. A comparison of MB-1 with other known B cell antigens suggests that the MB-1 antigen has not been previously identified. From one-and two-color immunofluorescence studies, it appears that the MB-1 antigen is found on all normal immunoglobulin (Ig)-expressing cells, but not on T cells, thymocytes, granulocytes, or platelets. Studies of malignant B cell tumors reveal that the antigen is expressed by virtually all Ig-expressing B cell tumors but only 10% of SIg- B-lineage leukemias. Data from these studies suggest that the MB-1 antigen is expressed late in B cell ontogeny before the expression of SIg.

Immunological phenotype of neoplasms involving the B cell in the last step of differentiation.

The immunological phenotype of diseases involving the last step of B cell differentiation--multiple myeloma (MM, 38 patients) and Waldenström's macroglobulinaemia (WM, 12 patients)--was analysed with a panel of monoclonal antibodies (McAb) as well as conventional markers. Most of the bone marrow plasma cells (80%) from MM patients reacted with the McAb OKT10, FMC8 and FMC48. Plasma cells were consistently negative with FMC7, Leu-1 and mouse rosettes. Ia, B1 and SIg were expressed in a minority of plasma cells (less than 30%) in half of the cases. The circulating neoplastic cells from five MM patients showed a more immature phenotype, with a higher reactivity for OKIa, B1 and increased SIg and a lower expression of CIg, than bone marrow plasma cells. The malignant cells of WM patients differed from those of MM in the reactivity with FMC7, being positive in 10 out of 11 cases, and in their high expression of B1, Ia and SIg with a predominant mu+ phenotype. Mouse rosettes and Leu-1 were positive in one case; OKT10 was positive in three out of five WM patients studied. This phenotype indicates that WM cells correspond to an earlier stage of B cell differentiation than MM plasma cells. The McAb J5 was positive in three out of six MM and two out of four WM analysed. The antigenic differences observed in MM and WM patients support the notion that the cells of the neoplastic clone are able to undergo a certain degree of differentiation.

Immunologic heterogeneity of diffuse large cell lymphoma

The cellular lineage of 57 diffuse large-cell lymphomas (DLCLs) was determined using a panel of monoclonal antibodies directed against lineage-restricted and -associated T, B, and monocyte antigens. The majority (82%) were of B cell lineage as determined by the expression of sig and/or B1, with the remaining 16% being of T cell lineage and 2%, of monocyte-myeloid lineage. By the expression of other B cell- restricted and -associated antigens, two major and two minor subgroups could be identified. These subgroups expressed the following phenotypes: (1) B1+B4+sIG+B2- (51%); (2) B1+B4+sIg+B2+ (29%); (3) B1+B4+sIg-B2+ (10%); and (4) B1+B4-sIg+B2- (10)%. The morphology of transformed lymphocytes, the weak to absent expression of the early B cell antigens B2 and sIgD, and the absence of the late B cell differentiation antigens PCA-1 and PC-1 suggested that these tumors were the neoplastic counterparts of normal B cells at the mid-stages of differentiation. Further support for the notion that B-DLCLs correspond to transformed B lymphocytes was concluded from the observation that B cells could be identified in normal spleen that expressed the cell surface phenotype and morphological appearance of the majority of B- DLCLs.

Immunologic Heterogeneity of Diffuse Large Cell Lymphoma

The cellular lineage of 57 diffuse large-cell lymphomas (DLCLs) was determined using a panel of monoclonal antibodies directed against lineage-restricted and -associated T, B, and monocyte antigens. The majority (82%) were of B cell lineage as determined by the expression of sig and/or B1, with the remaining 16% being of T cell lineage and 2%, of monocyte-myeloid lineage. By the expression of other B cell-restricted and -associated antigens, two major and two minor subgroups could be identified. These subgroups expressed the following phenotypes: (1) B1+B4+sIG+B2- (51%); (2) B1+B4+sIg+B2+ (29%); (3) B1+B4+sIg-B2+ (10%); and (4) B1+B4-sIg+B2- (10)%. The morphology of transformed lymphocytes, the weak to absent expression of the early B cell antigens B2 and sIgD, and the absence of the late B cell differentiation antigens PCA-1 and PC-1 suggested that these tumors were the neoplastic counterparts of normal B cells at the mid-stages of differentiation. Further support for the notion that B-DLCLs correspond to transformed B lymphocytes was concluded from the observation that B cells could be identified in normal spleen that expressed the cell surface phenotype and morphological appearance of the majority of B-DLCLs.