IL-4, but not IL-5, can act synergistically with B cell activating factor (BCAF) to induce proliferation of resting B cells

B cell activating factor (BCAF) was initially identified in the supernatant of the murine T helper cell clone 52-3 (52-3 SN) because of its ability to promote activation and proliferation of resting B cells in the absence of any other costimulus. In this paper, we show that 52-3 T helper cells also secrete IL-4 and IL-5 and we have analyzed the influence of these two lymphokines on B cell proliferation induced by BCAF-containing 52-3 SN. Using the neutralizing anti-IL-4 monoclonal antibody 11B11, we observed partial inhibition of B cell proliferation. 52-3 SN free of IL-4 prepared using an immunoabsorbent column was still able to induce significant B cell proliferation. Although recombinant IL-4 alone does not induce B cell proliferation, it increased the proliferation induced by IL-4-free 52-3 SN. Kinetic studies showed that IL-4 is required at the start of B cell cultures in order to exert optimal synergistic effects. In contrast, anti-IL-5 monoclonal antibody NC17 did not affect the B cell proliferative activity of 52-3 SN whether or not IL-4 was present. When 52-3 SN was tested on dextran-sulfate-activated B cells, IL-5 and BCAF activities were detected but only the IL-5 activity was neutralized by monoclonal antibody NC17. These results demonstrate that (i) BCAF-containing SN can induce proliferation of resting B cells independently of IL-4 and IL-5, and (ii) IL-4, but not IL-5, can act synergistically with BCAF to induce B cell proliferation.     

Regulation of IgA differentiation in CH12LX B cells by lymphokines. IL-4 induces membrane IgM-positive CH12LX cells to express membrane IgA and IL-5 induces membrane IgA-positive CH12LX cells to secrete IgA

In these studies we utilized the Ag (SRBC)-reactive B cell line CH12LX to study isotype switching. CH12LX cells are a stable population of B cells mainly bearing membrane IgM (mIgM) (98 to 99%) with a small population of B cells bearing membrane IgA (mIgA) (1 to 2%). LPS induced a 5- to 10-fold increase in the secretion of both Ig, whereas a lymphokine-rich supernatant of D10 T cells induced a greater increase in the secretion of IgA than IgM. Analysis of the latter effect with recombinant lymphokines disclosed that rIL-4 induced an increase in the number of mIgA+ cells (6 to 15%) with minimal effect on IgA secretion, whereas IL-5 induced increased IgA secretion but had no effect on mIgA expression. The addition of both lymphokines induced increased mIgA expression and IgA secretion. No effect on mIgA expression or IgA secretion was seen with other lymphokines, including IL-1, IL-2, IL-3, IL-6, GM-CSF, and IFN-gamma. The rIL-4 effect on CH12LX cells represents true differentiation rather than selective proliferation for the following reasons: first, subclones of CH12LX cells respond to IL-4-containing T cell supernatant in the same fashion as the original cell line; second, culture of CH12LX cells with IL-4 causes the appearance of large numbers of dual-bearing mIgM/mIgA cells as well as mIgA+ cells and a dual-bearing mIgM/mIgA line was obtained by cloning CH12LX after stimulation with an IL-4-containing supernatant; third, sorted mIgA+ and mIgA- CH12LX cells had similar rates of proliferation in the presence or absence of IL-4. In further studies, it was found that IL-5 causes IgA secretion by mIgA+ but not mIgA- CH12LX cells indicating that it is acting as a post-isotype switch differentiation factor. These studies are consistent with the view that IL-4 and IL-5 act in a sequential fashion to induce IgA expression and secretion in CH12LX cells, IL-4 inducing differentiation of mIgM+ cells to mIgA+ cells and IL-5 enhancing the IgA secretion by the resulting mIgA-bearing cells.     

Both B151-T cell replacing factor 1 and IL-5 regulate Ig secretion and IL-2 receptor expression on a cloned B lymphoma line

In the present study, we have demonstrated that both B151-T cell-replacing factor 1 and rIL-5 are responsible for the activity to partially induce CL-3 cells into IgM-synthesizing cells and also to synergize with IL-2 to augment IL-2R expression on and IgM synthesis in CL-3 cells. These actions of rIL-5 on a homogeneous cloned line (BCL1-CL-3 cells) allow us to identify and characterize the two alternated B cell developmental pathways. One is an IL-2-independent, IL-5-driven differentiation pathway without preceding up-regulated IL-2R expression, and the other is an IL-5 plus IL-2-dependent augmented differentiation pathway with preceding up-regulated IL-2R expression. We have also demonstrated the functional difference of two distinct B cell growth-promoting factors, B cell-stimulating factor 1 (rIL-4) and rIL-5. CL-3 cells are equally stimulated to grow by rIL-4 and rIL-5, whereas only rIL-5 can render CL-3 cells responsive to rIL-2, indicating that these two lymphokines affect B cells in a strikingly different manner.     

Growth regulation of multi-factor-dependent myeloid cell lines: IL-4, TGF-beta and pertussis toxin modulate IL-3- or GM-CSF-induced growth by controlling cell cycle length

The stimulatory effects of lymphokines, interleukin 3 (IL-3), granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin 4 (IL-4), and the inhibitory effects of transforming growth factor beta (TGF-beta) and the pertussis toxin, islet activating protein (LAP), on multi-factor-dependent myeloid cell lines were examined. The effects of IL-3 on a mast cell progenitor clone, IC2 were indistinguishable from those of GM-CSF with respect to their concentration-response curves for induction of DNA synthesis and capability to maintain cell growth for many months. IL-4 acts differently on IC2 cells: the maximum level of DNA synthesis induced by IL-4 is always lower than that induced by IL-3 or GM-CSF and IL-4-induced proliferation is transient. IL-4, however, synergistically induced DNA synthesis of IC2 cells with limiting concentrations of IL-3 or GM-CSF. When IC2 cells were cultured with saturating concentrations of IL-3, GM-CSF or a combination of both, the doubling time was 25 +/- 1 h, whereas it decreased to 17 +/- 1 h when IL-4 was further added to the cultures. IAP reduced the DNA synthesis of IC2 cells induced by the above three growth factors. The doubling time of IC2 cells was 30 +/- 2 h when IC2 cells were cultured with sufficient concentrations of IL-3 in the presence of IAP. Cell cycle analysis revealed that the fraction of cells in Gl was decreased by IL-4 but was increased by IAP. TGF-beta also reduced IL-3-dependent DNA synthesis and increased the fraction of cells in Gl. The inhibitory effect on IL-3-dependent growth of IC2 cells was not increased when these cells were exposed simultaneously to TGF-beta and IAP. The results suggest that IL-3 and GM-CSF stimulate the growth of IC2 cells through similar pathways and that IL-4 augments the action of IL-3 or GM-CSF by decreasing the Gl period. It is also suggested that IAP and TGF-beta retard the growth of IC2 cells by increasing the fraction of cells in GI.     

Detection and characterization of a B cell stimulatory factor (BSF-TC) derived from a bone marrow stromal cell line

Stromal cell lines derived from murine bone marrow support the growth of immature pre-B cells and produce cytokines that affect the growth and differentiation of other hematopoietic precursors. Conditioned medium (CM) from one such line (TC-1) stimulated marked proliferation of B cells previously activated by anti-Ig (anti-Ig blasts). Proliferation of anti-Ig blasts was not induced by purified cytokines known to be produced by TC-1 (CSF-1, GM-CSF, or G-CSF) or by IL-1, IL-2, IL-3, IL-4, IL-5, or IL-6. Furthermore, IL-2, IL-4, and IL-5, alone or in combination, failed to support proliferation or differentiation of anti-Ig blasts. TC-1 CM enhanced proliferation of B cells that were co-cultured with LPS, anti-Ig, or dextran sulfate; co-stimulation with anti-Ig was unaffected by the presence of monoclonal anti-IL-4. Proliferation of low, but not high, density B cells isolated from spleen was directly stimulated by TC-1 CM. These results suggest that bone marrow stromal cells produce a novel B cell stimulatory factor (BSF-TC) that induces proliferation of activated B cells.     

Enhancement of IL-2-induced T cell proliferation by a novel factor(s) present in murine spleen dendritic cell-T cell culture supernatants

The mechanism(s) underlying the potent accessory cell function of dendritic cells (DC) remains unclear. The possibility was considered that a soluble factor(s) released during the interaction of DC and T cells might contribute to the potent T cell activating function of DC. Culture supernatants were generated from mixtures of murine spleen DC and periodate-treated spleen T cells and were examined for the presence of known cytokine activities and factors capable of enhancing T cell responsiveness to IL-2. Serum-free supernatants from 24 h DC-T cell co-cultures exhibited high levels of IL-2, detectable levels of IL-3, and negligible levels of IL-1, -4, -5, -6, and TNF. A factor(s) was also identified with an apparent Mr of 12.5 to 17.0 kDa, henceforth designated IL-2 enhancing factor (IL-2EF), which enhanced the IL-2-induced proliferation of murine thymocytes, CTLL, and HT-2 cells by approximately three- to fourfold. This enhancement was also observed in the presence of neutralizing antibodies to murine IL-1 alpha, -1 beta, -3, -4, -5, -6, granulocyte-macrophage (GM)-CSF, TNF, and IFN-gamma. However, IL-2EF failed to enhance: 1) the activity of IL-1, -3, -4, -5, or -6 on cells responsive to these cytokines; 2) IL-2-augmented, IL-5-induced BCL1 proliferation; and 3) either PHA- or Con A-stimulated thymocyte proliferation. Moreover, neither IFN-gamma nor GM-CSF exhibited IL-2EF activity. When DC and T cells were cultured separately (after an initial 12 h co-culture period), IL-2EF activity resided predominantly in the T cell-derived supernatants. These and other data indicate that IL-2EF, a heat-labile T cell-derived 12.5 to 17.0 kDa protein, is distinct from IL-1 alpha, -1 beta, -2, -3, -4, -5, -6, TNF, IFN-gamma, GM-CSF, and previously described factors that co-stimulate thymocyte proliferation in the presence of Con A or PHA. It is suggested that IL-2EF functions to specifically enhance IL-2-driven T cell proliferation and contributes to the potent activation of T cells induced by DC.     

IL-3, IL-4, and IL-6 enhance IFN-gamma-dependent bone marrow natural suppressor activity

The ability of murine bone marrow (BM) natural suppressor (NS) cells to suppress a Con A proliferation assay was greatly enhanced by supernatant obtained from the T cell hybridoma D9C1.12.17. Of the lymphokines produced by this hybridoma, three were found to enhance suppression: interleukin-3 (IL-3), IL-4, and IL-6. These molecules enhanced suppression of both unirradiated and irradiated (2000 R) BM cells indicating that augmented suppression was not just due to proliferation of NS cells. The ability of all three of the lymphokines to enhance BM suppression could be blocked by anti-interferon-gamma (IFN-gamma) antibody. These results indicate that (1) NS cell activity is not radiosensitive and (2) that two signals may be required for maximal NS cell suppression, one being a lymphokine-mediated signal and the other IFN-gamma.     

Effects of hematopoietic growth factor (GM-CSF: granulocyte-macrophage colony-stimulating factor) on thymocyte proliferation induced by interleukin-1 (IL-1)

A semi-purified fraction obtained from P388 D1 cell line conditioned medium (P388 D1 CM) which contains Interleukin-1 (IL-1) and Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) stimulates murine thymocyte proliferation both in the absence and the presence of a suboptimal dose of phytohemagglutinin (PHA). Because this effect on thymocyte proliferation is always larger than that obtained with optimal concentrations of pure IL-1, we have investigated the possible involvement of GM-CSF in this semi-purified fraction mediated-thymocyte proliferation. We here show that the maximal level of thymocyte proliferation induced by the semi-purified fraction is comparable to that obtained by the co-addition of recombinant GM-CSF and IL-1. In addition, although GM-CSF alone induces no significant thymocyte proliferation, the presence of an anti-GM-CSF antiserum partially blocks the thymocyte proliferation induced by the semi-purified fraction. Thus, the capacity of the semi-purified fraction of P388 D1 to stimulate thymocyte proliferation appears to result from a synergistic action between GM-CSF and IL-1.     

Tumor necrosis factor-alpha as a proliferative signal for an IL-2-dependent T cell line: strict species specificity of action

The ability of TNF-alpha to stimulate T cell proliferation was examined. We demonstrate that murine rTNF-alpha induces the proliferation of CT6, a murine T cell line previously thought to be responsive only to IL-2. This activity appears to be the result of the direct action of murine rTNF-alpha on the CT6 cells because neither 1) murine IL-2 or murine IL-4, lymphokines also capable of inducing CT6 proliferation, were detected in culture supernatants from murine rTNF-alpha-treated CT6 cells nor 2) did antibodies specific for IL-2 or IL-4 inhibit murine rTNF-alpha-induced CT6 proliferation. Unlike many of the activities displayed by TNF-alpha, its ability to induce CT6 cell proliferation shows strict species specificity as indicated by the failure of human rTNF-alpha to stimulate these cells. Flow cytometric analysis and binding of radiolabeled TNF-alpha have indicated that receptors for TNF-alpha on these cells are specific for murine TNF-alpha. The ability of murine rTNF-alpha to induce the proliferation of certain T cell lines further indicates that this molecule plays an important role in regulation of T cell-mediated immune responses.     

IL-4 regulates differentiation and proliferation of human precursor B cells

The mechanism by which precursor and pre-B cells undergo differentiation is unclear; however, it is known that growth factors play an important role in this maturation process. The lymphokine, IL-4 has been shown to increase expression of class II Ag on B cells and induce B cell proliferation. In the murine system, IL-4 induced differentiation of precursor B cells into pre-B cells. In order to analyze growth factors on B cell development we have established an in vitro culture system for human bone marrow cells. We found that in the presence of IL-4, normal human precursor and pre-B cells can be induced to differentiate in the absence of cell proliferation with four days of culture. Furthermore, IL-4 depressed proliferation induced by supernatant from a T cell line. The differentiation was measured by an increase in both the number of cytoplasmic mu and surface IgM-positive cells. The effect of IL-4 on precursor and pre-B cell differentiation was detected as soon as 14 h of exposure to the lymphokine in the absence of an adherent feeder layer. These data suggest that IL-4 directly affects the differentiation process of normal human precursor and pre-B cells, and may antagonistically affect cell proliferation.     

Characterization of a subclone (D10S) of the D10.G4.1 helper t-cell line which proliferates to attomolar concentrations of interleukin-1 in the absence of mitogens

Most studies have shown that interleukin-1 (IL-1) acts as a helper or co-stimulator in T-lymphocyte activation and proliferation by mitogens or antigens. We describe here a stable subclone (D10S) of the murine D10.G4.1 helper T-cell which proliferates to subfemtomolar (attomolar) concentrations of IL-1 beta or alpha in the absence of mitogens. D10S cells have been maintained in culture for over two years without splenic cell feeder layers nor antigen stimulation. Detection of proliferation can be made by either uptake of tritiated thymidine at 72 h or in 48 h by a colorimetric assay which measures mitochondrial dehydrogenases; the latter assay is rapid and inexpensive. D10S cells are distinct from the parent clone D10.G4., which requires mitogens for IL-1 activity. IL-1-induced proliferation is independent of the elaboration of IL-2, IL-4, or IL-6, although these cells proliferate to these lymphokines at considerably higher concentrations when compared to IL-1. The D10S cells proliferate in direct correlation to the duration of IL-1 presence in the culture. We found no evidence that IL-1 induced more IL-1 in these cells. The subclone is highly specific for IL-1: proliferation was not observed to endotoxin, human or murine interferon-gamma (IFN gamma), tumor necrosis factor (TNF), lymphotoxin, or granulocyte-macrophage colony stimulating factor (GM-CSF). There was no suppressive effect of transforming growth factor (TGF beta). Only at high concentrations (100 ng/ml) did IL-6 induce proliferation. We conclude that this stable, feeder layer-free cell line is highly sensitive to IL-1 which acts as a direct stimulant for these cells; they are also useful for bioassays as well as the study of IL-1 receptors as described in the accompanying paper.     

Interleukin-5 (IL-5) and IL-6 define two molecularly distinct pathways of B-cell differentiation.

Interleukin-5 (IL-5) and IL-6 have both been reported to act as B-cell differentiation factors by stimulating activated B cells to secrete antibody. However, it has not been possible to directly compare the effects of these two lymphokines because of the lack of a suitable B-cell line capable of responding to both. We have identified a clonal, inducible B-cell lymphoma, CH12, that has this property. Both IL-5 and IL-6 can independently stimulate increases in steady-state levels of immunoglobulin and J-chain mRNA and proteins, and they both induce the differentiation of CH12 into high-rate antibody-secreting cells. Nevertheless, there are significant differences in the activities of these two lymphokines. First, while IL-6 acts only as a differentiation factor, IL-5 also augments the proliferation of CH12 cells. Second, the differentiation stimulated by IL-5 but not by IL-6 is partially inhibited by IL-4. Inhibition of IL-5-induced differentiation was not at the level of IL-5 receptor expression, since IL-4 did not inhibit IL-5-induced proliferation. Third, IL-5 but not IL-6 stimulated increased mouse mammary tumor proviral gene expression in CH12 cells. These results demonstrate that while both IL-5 and IL-6 may act as differentiation factors for B cells, they induce differentiation by using at least partially distinct molecular pathways. Our results also establish that B cells characteristic of a single stage of development can independently respond to IL-4, IL-5, and IL-6.     

Establishment and characterization of a unique human cell line that proliferates dependently on GM-CSF, IL-3, or erythropoietin

We have established a novel cell line, designated as TF-1, from a patient with erythroleukemia, which showed complete growth dependency on granulocyte-macrophage colony-stimulating factor (GM-CSF) or on interleukin-3 (IL-3) and carried a homogeneous chromosomal abnormality (54X). Erythropoietin (EPO) also sustained the short-term growth of TF-1, but did not induce erythroid differentiation. These three hematopoietic growth factors acted on TF-1 synergistically. Transforming growth factor-beta and interferons inhibited the factor-dependent growth of TF-1 cells in a dose-dependent fashion, and monocyte-colony stimulating factor and interkeukin-1 enhanced the GM-CSF-dependent growth of TF-1. Ultrastructural studies revealed some very immature features in this cell line. Although TF-1 cells do not express glycophorin A or carbonyl anhydrase I, the morphological and cytochemical features, and the constitutive expression of globin genes, indicate the commitment of TF-1 to erythroid lineage. When induced to differentiate, TF-1 entered two different pathways. Specifically, hemin and delta-aminolevulinic acid induced hemoglobin synthesis, whereas TPA induced dramatic differentiation of TF-1 into macrophage-like cells. In summary, TF-1 is a cell line of immature erythroid origin that requires GM-CSF, IL-3, or EPO for its growth and that has the ability to undergo differentiation into either more mature erythroid cells or into macrophage-like cells. TF-1 is a useful tool for analyzing the human receptors for IL-3, GM-CSF, and EPO or the signal transduction of these hemopoietic growth factors.     

IL-6 is a potent cofactor of IL-1 in IgM synthesis and of IL-5 in IgA synthesis

In these studies we determined the capacity of IL-6 to act as a differentiation cofactor for murine Peyer's patch B cells producing different Ig classes and subclasses. In preliminary studies we determined that sufficient endogenous IL-6 was produced in LPS-induced cell systems to obscure responses to exogenous IL-6. We therefore studied IL-6 effects on Peyer's patch B cells (T cell-depleted cell populations) in the absence of LPS, relying on responses of in vivo-activated cells. rIL-1 alpha or purified IL-6 only slightly enhanced synthesis of IgM over minimal baseline levels in Peyer's patch T cell-depleted cell cultures; however, when IL-6 was added to cultures also containing rIL-1, IgM synthesis was very substantially increased. In addition, rIL-5 alone gave rise to a modest increase in IgM synthesis and its effect was not enhanced by either rIL-1 or IL-6. IgG production (mainly IgG3) followed a similar pattern. In contrast, IgA production was only modestly increased above baseline by rIL-1, rIL-5, or IL-6 alone or by rIL-1 and IL-6 in combination, but was greatly increased by rIL-5 and IL-6 in combination. The effect of IL-6 on Ig synthesis in the above studies was not due to an effect on cell proliferation. In summary, these data indicate that B cells differ in respect to the cytokines supporting maximal terminal differentiation and thus the class of Ig produced may depend on the presence of a particular combination of cytokines and lymphokines.     

Prostaglandin E2-dependent induction of granulocyte-macrophage colony-stimulating factor secretion by cloned murine helper T cells

PG are known to inhibit T cell proliferation, at least in part by suppressing IL-2 production, but effects of PG on the production of other lymphokines have not been well studied. We have found that PGE2 and PGE1, but not PGF2 alpha, inhibit both proliferation and production of granulocyte-macrophage (GM)-CSF by murine TH clones stimulated with Ag or anti-CD3 antibody. Thus, signals generated via the Ag receptor:CD3 complex were inhibited by PGE. Most interesting, however, was the finding that PGE2 and PGE1 could act synergistically with IL-2 for the induction of GM-CSF in some TH1 clones. Dependence on PGE2 for this response was not found in all clones, as some TH1 cells could produce GM-CSF after IL-2 alone, and some cells did not produce GM-CSF even in the presence of PGE2 and IL-2. These observations indicate that there is a subset of TH1 cells receptive to a stimulating activity of PGE2 in the presence of IL-2. PGE2 is known to elevate cAMP levels in T cells. Therefore, we tested whether other agents known to increase cAMP, such as forskolin and cholera toxin, could act in conjunction with IL-2 to induce GM-CSF secretion. As was found with PGE2, these compounds also induced GM-CSF activity in the presence of IL-2, suggesting a critical role for cAMP in this process. Overall these data indicate that the requirements for activation of GM-CSF secretion vary among individual T cells. Most importantly they provide the first evidence that E-series PG are positive signals for lymphokine induction in certain T cells, whereas simultaneously acting as negative signals limiting proliferation. This result also suggests that treatment with anti-inflammatory drugs that decrease PGE2 concentrations may inhibit lymphokine secretion normally stimulated by this pathway.     

白细胞介素—2加强小鼠T淋巴细胞产生白细胞介素—3

In addition to the regulation of T cell growth, IL-2 exerts effects on the induction of certain lymphokines. We show here that IL-2 synergizes with 5 micrograms/ml of ConA to promote the production of IL-3 in mouse splenic T cell cultures. IL-3 was measured as CFU-GEMM-inducing activity on mouse bone marrow progenitor cells in the supernatant of the stimulated mouse splenic T cells (TCM). The resting T cells produced no CFU-GEMM-inducing activity, but could be induced to produce low level of CFU-GEMM-inducing activity in the presence of ConA. In vitro exposure to IL-2 markedly increased CFU-GEMM-inducing activity production (nearly up to 8-fold) by the ConA-activated T cells. Optimal stimulation was observed when 80 u/ml IL-2 was used for 48 h incubation. Anti-mouse IL-3 monoclonal antibody inhibited the CFU-GEMM inducing activity of TCM. Moreover, the TCM stimulated the proliferation of IL-3 dependent cell line FDC-P1. We also show that IL-2 and ConA-treated T cells expressed high level of IL-3 mRNA through dot blot analysis. These results confirmed the nature of CFU-GEMM-inducing activity of TCM as IL-3. The capacity of IL-2 to promote the production of IL-3 may represent an important mechanism by which it mediate the communication between the immune and hematopoietic systems.     

Interleukin 5 (IL-5) can act in G0/G1 to induce S phase entry in B lymphocytes from normal and autoimmune strain mice and in transformed leukemic B cells

In addition to its ability to enhance antibody secretion, Interleukin 5 (IL-5) enhances murine B lymphocyte proliferation. This so-called growth factor activity has been amply demonstrated by many laboratories assessing thymidine incorporation or cell recovery. Attempts to actually quantitate the fraction of fresh splenic B cells responding to IL-5, by limiting dilution analysis or other means, with few exceptions have yielded disappointingly small numbers--generally between 1 and 5%, or perhaps less. We have recently identified the peritoneal cavity as a reservoir rich in IL-5-responsive B cells. In this report, we provide independent corroboration of this high IL-5 reactivity by means of cell cycle analysis. Low-density peritoneal B cells, more than 90% of which are in G0 and G1 phases, were stimulated with polyclonal activators in the presence of mitotic inhibitors. Frequencies of IL-5-responsive B cells were measured by observing the differences in the proportions of cultured cells entering S and later phases in the presence, compared to the absence, of IL-5. Some 10 to 20% more of the low-density peritoneal B cells from normal mice entered S phase when IL-5 was present with LPS + DXS. A similar IL-5-mediated elevation in the frequency of S phase entry was seen with peritoneal B cells from the autoimmune mouse strain NZB. Furthermore, a measurable fraction of peritoneal B cells from these mice were even capable of responding to IL-5 alone. These IL-5-induced increases could be blocked by anti-IL-5 mAb. About 30% of the BCL1 leukemic B cell line initiated DNA replication when stimulated with IL-5 alone. Hence, IL-5-responsive B cell fractions have been measured for some normal, autoimmune strain and transformed leukemic B cell phenotypes. In addition to quantitating the proportion of IL-5-responsive B cells, these experiments formally demonstrate that IL-5 can act in the G1 phase to increase S phase entry.     

Human B cell proliferation in response to IL-4 is associated with enhanced production of B cell-derived growth factors

To investigate the capacity of human IL-4 to function as a B cell growth factor (BCGF), we studied its ability to promote proliferation of a selected B cell line. We show that the cell line, designated A4, proliferated in response to IL-4 in a dose-dependent manner. The A4 cells also proliferated in response to their own B cell derived growth factor (B. BCGF), suggesting autocrine-mediated growth. The ability of IL-4 to induce proliferation of the A4 cell line was dependent on the level of autocrine growth. At low cell density, IL-4 induced marked dose-dependent proliferation. However, as A4 cell density increased, the ability of IL-4 to induce proliferation was diminished. The possibility that IL-4 may be mediating the autocrine growth of A4 cells was ruled out, because A4 cell-derived BCGF failed to induce CD23/low affinity receptors for the Fc region of IgE on activated tonsillar B cells and anti-IL-4 antibody did not block B. BCGF activity. We found that IL-4 stimulation of A4 cells and activated tonsillar B cells is associated with enhanced production of B. BCGF. These data indicate that human IL-4 has the capacity to promote proliferation of the B cell line A4, and that the ability of IL-4 to function as BCGF is associated with enhanced autocrine growth of activated B cells.     

Cognate interactions between helper T cells and B cells. V. Reconstitution of T helper cell function using purified plasma membranes from activated Th1 and Th2 T helper cells and lymphokines.

Th physically interact with B cells and produce lymphokines that influence B cell growth and differentiation. The respective contribution of cell contact and lymphokines to induction of B cell growth and differentiation was addressed using purified plasma membranes (PM) from resting Th (PMrest) and anti-CD3-activated Th (PMCD3) together with lymphokines. Results show that PMCD3, but not PMrest, induce 10% of resting B cells to enter the G1 phase of the cell cycle, with few B cells entering G1b and S/G2. The inclusion of IL-4, but not IL-2, IL-5, or IFN-gamma, amplifies the B cell response to PMCD3 by increasing the total percentage of activatable B cells to greater than 40% and inducing B cell progression into G1b, S, and G2. Direct comparison between PMrest and PMCD3 purified from Th1 and Th2 indicate that both Th1 and Th2 induce similar levels of B cell proliferation in the presence of IL-4. Further, the lymphokine requirements for B cell proliferation induced by PMCD3 from Th1 and Th2 is indistinguishable. B cell differentiation to IgM, IgG1, and IgG2a synthesis by PMCD3 required IL-4 and IL-5. Using lymphokine conditions that supported B cell differentiation, PMCD3 purified from Th1 and Th2 induced similar levels of IgM, and IgG1. Given the functional data on PMCD3 from Th1 and Th2, the data indicate that there are no substantive differences between Th1- and Th2-derived PMCD3, and that the major differences in the ability of viable Th1 and Th2 to activate B cells is the lymphokines produced by the cells.