Klaus Beiske

Transforming growth factor type β (TGF β) inhibits G1 to S transition, but not activation of human B lymphocytes☆

Abstract Type β transforming growth factor (TGF β) is a polypeptide that may influence the growth of a variety of cell types in a positive or negative fashion. In this study we show that TGF β markedly inhibits DNA synthesis in normal and neoplastic human B lymphocytes stimulated to proliferate with anti-immunoglobulins and B-cell growth factor (BCGF). Although TGF β was needed during the initial 12 h of the culture to promote optimal inhibition, we found that it had little or no effect on several early to intermediate parameters of cell activation ([Ca 2+ ] i increase, c- myc mRNA increase, cellular enlargement, RNA increase, and the increase in the expression of the 4F2 activation antigen). In contrast, TGF β almost completely blocked the induction of transferrin receptor expression, which normally occurs in the late G 1 phase of the cell cycle. Therefore, we conclude that TGF β treatment leads to arrest of the cells in the middle to late G 1 phase, prior to transferrin receptor expression.

Immunological typing of acute leukemias: Immunoenzymatic staining of fixed cells compared with immunofluorescence staining of unfixed cells in suspension

A panel of 14 monoclonal antibodies (McAb) against hematopoietic cell surface antigens was applied on mononuclear blood or bone marrow cells from 40 cases of acute leukemia in order to compare immunoenzymatic staining (IE) (alkaline phosphatase) of fixed cells with immunofluorescence staining (IF) of unfixed suspended cells. According to the immunological results, 25 cases were phenotyped as ALL and 15 cases as AML. Cases with blast crisis secondary to chronic myelogeneous leukemia (CML‐BC) were not represented in this series. In all ALL cases the two methods gave an identical antigenic distribution. In 20 our of 21 cases of non‐T‐cell ALL, a B‐cell progenitor origin was demonstrated by a positive staining reaction with the anti‐CD 19 McAb AB1 or HD37, and in 10 cases additionally with the anti‐CD20 McAb B1 or 1F5. In contrast to the results obtained with IF, IE revealed a poor preservation of the AB1 epitope on CD19, whereas the HD37 epitope was equally well demonstrated by both methods. In 15 cases of AML the distribution of positive versus negative cells with IE or IF was identical for all McAb except J5 (anti‐CALLA) (CD10) and Bl (CD20). Thus, 10/15 AML cases expressed CALLA with IE compared to 2/15 with IF. The corresponding figures for Bl were 5/15 and 0/15, respectively. Accordingly, normal myeloid precursor cells were CALLA‐positive with IE but negative with IF. The discrepancy probably reflects the fact that, whereas both intracytoplasmatic and membrane‐bound antigens are exposed in IE, only the latter are in IF. If the alteration of antigenic accessibility after fixation is considered, IE can safely be used for immunophenotyping of acute leukemia.

Different responses elicited in vitro by antibodies to IgM and IgD in cells from a surface IgM + D-positive human follicular lymphoma.

The effect of anti‐Igs in combination with the tumour promoter TPA on DNA and Ig synthesis in a human follicular germinal centre cell lymphoma carrying slgM and slgD was investigated. The lymphoma cells responded to both amiti‐μ and anti‐γ chains and to anti‐δ light chains by DNA synthesis, as measured by mcthy‐(3H)‐thymidine incorporation. Flow cytofluorometric measurements, however, showed that only anti‐μ chains induced marked increase in cytoplasmic Ig content. This result suggests that in certain B‐cell subsets the signals elicited via slgM and slgD are different.

Induction of Maturation in Different Types of B‐Cell Lymphomas in Vitro with TPA and Antibodies to Surface Immunoglobulin

Cells from eight selected cases of human non‐Hodgkin lymphomas of various histological types (lymphocytic, centrocytic, centrocytic/centroblastic, and immunocytomas) were stimulated in vitro with 12‐O‐tetradecanoyl‐phorbol‐13‐acetate (TPA) and anti‐immunoglobulins (anti‐Ig) against the surface immunoglobulin (sIg) on the tumour cells. Six of these cases responded by intracellular Ig accumulation as measured by flow cytofluorometry and direct phenotypical change into immunoblasts/plasmablasts as detected by light microscopic immunocytochemistry. However, the response to TPA alone varied considerably from ease to case. These findings suggest that many, if not all, B‐cell subsets have the capacity to develop directly into Ig‐synthesizing cells (immunoblasts and plasmablasts). However, conditions for eliciting such events may vary, depending on the phenotypical properties and differentiation stage.

Regulation of c-myc mRNA and Protein Levels During Activation of Normal Human B Cells

The myc oncogene has been implicated in the control of cellular proliferation in both normal and neoplastic cells. Recently, several cell lines have been shown to express c-myc mRNA and protein throughout the cell cycle (Thompson et al 1985; Hann et al 1985; Rabbitts et al 1985). However, in several cell systems triggering of quiescent cells into G1 is accompanied by a marked burst of c-myc mRNA, peaking a few hours after stimulation (Kelly et al 1983; Goustin et al 1985; Smeland et al 1985a). This has focused on a possible function for the c-myc product during the induction of competence to respond to progression growth factors acting in G1. This theory finds support in studies on fibroblast cell lines, where experimental manipulation of c-myc levels was related to cell cycle progression (Armelin et al 1984; Kaczmarek et al 1985). In both cases the increased myc levels provoked an increased sensitivity to the action of progression factors.

Different Fate of Antibodies to Surface IgM and IgD in Germinal Centre Cell-Associated Lymphomas

In all of four human germinal centre cell‐associated B lymphomas carrying sIgM + sIgD F(ab)2 fragments of rabbit antibodies to human μ‐chain were accumulated intracellularly, whereas the accumulation of antibodies to δ‐chain was considerably less abundant. The accumulation of antibodies to light chain was intermediate between antibodies to μ‐ and δ‐chain. These results could be explained by the following observations: (i) the difference in the level of reexpression of sIgM and sIgD and (ii) the discovery that antibodies to δ‐chain bound to reexpressed sIgD were degraded at a higher rate than those bound to primary sIgD and at a higher rate than antibodies to μ‐chain, whether hound to primary or reexpressed sIgM.