Chihiro Ishioka

Human recombinant erythropoietin directly stimulates B cell immunoglobulin production and proliferation in serum‐free medium

The effect of human recombinant erythropoietin (Epo) on B cell responses was studied in a serum‐free medium. Epo enhanced IgM production and thymidine uptake by a human IgM‐producing lymphoblasloid cell line, CBL. This effect was specific to Epo since enhancement was blocked by anti‐Epo antibody but not by control antibody. Among the various cytokines, interleukin‐4 (IL‐4) enhanced IgM production and thymidine uptake while IL‐6 enhanced IgM production without affecting thymidine uptake. In contrast, other cytokines including IL‐1β, IL‐2, IL‐5, interferon‐alpha (IFN‐α), interferon‐gamma (IFN‐γ), or granulocyte/macrophage colony‐stimulating factor (GM‐CSF) were without effect. However, the enhancing effect of Epo is different from that of IL‐4 or IL‐6, since Epo effect was not blocked by anti‐IL‐4 antibody or anti‐IL‐6 antibody. Moreover, specific binding of Epo was detected on CBL cells. Epo also enhanced immunoglobulin (IgG, IgM and IgA) production and thymidine uptake by purified tonsil small resting B cells stimulated by Staphylococcus aureus Cowan strain I (SAC) or by large activated B cells. In contrast, Epo had no effect on unstimulated smalt resting B cells. These results indicate that Epo could directly stimulate activated and differentiated B cells and could enhance B cell immunoglobulin production and proliferation.

Vasoactive intestinal peptide stimulates immunoglobulin production and growth of human B cells

The effect of vasoactive intestinal peptide (VIP) on human lymphoblastoid B cell lines and tonsil B cells was studied. VIP increased immunoglobulin production and proliferation by lymphoblastoid B cell line. GM‐1056, in a dose‐dependent manner. As little as 10‐12 M of VIP was effective, and higher concentrations of VIP induced an approximately five‐fold increase in IgA production. Moreover, this enhancement was blocked by VIP antagonist. Similarly, VIP enhanced IgM and IgG production by other lymphoblastoid B cell lines, CBL and IM‐9, respectively. In contrast to VIP, another neuropeptide substance P (SP) or somatostatin failed to enhance immunoglobulin production and thymidine uptake. VIP also enhanced IgA production and thymidine uptake by purified tonsil B cells. However, in contrast to B cell lines, VIP failed to enhance IgM and IgG production by tonsil B cells. SP or somatostatin failed to enhance immunoglobulin production or thymidine uptake by tonsil B cells. These results indicate that VIP acts as B cell stimulatory factor and that VIP may also have preferential effect on IgA production on tonsil B cells.

Differential effect of vasoactive intestinal peptide, somatostatin, and substance P on human IgE and IgG subclass production

We studied the effect of vasoactive intestinal peptide (VIP), somatostatin (SOM), and substance P (SP) on IL-4-stimulated human IgE and IgG subclass production. VIP and SOM, but not SP, inhibited IgE production without affecting IgM or IgA production by mononuclear cells (MNC) from nonatopic donors from 10 pM to 10 nM. These neuropeptides also differentially modulated IgG subclass production. While IgG1 production was not affected by VIP, SOM, or SP, all of the neuropeptides enhanced IgG2 production. By contrast, SOM and SP, but not VIP, inhibited IgG3 production, whereas VIP and SP, but not SOM, enhanced IgG4 production. The effect by neuropeptides was specific since each peptide effect was specifically blocked by each antagonist. To achieve this effect, neuropeptides must be added at the start of the culture and be present throughout the entire culture period. The inhibition of IgE production was not mediated by known inhibitors of IgE production, IFN-gamma or PGE2, because the addition of anti-IFN-gamma mAb (10 micrograms/ml) or indomethacin (0.1 microM) did not overcome the inhibition of IgE production. In contrast to MNC, neuropeptides did not affect IgG subclass production in purified B cells. IgE production was not induced by IL-4 in purified B cells. Neuropeptides also failed to modulate IgG subclass production in cultures of B cells with either T cells or monocytes. However, they modulated IgE production and IgG subclass production in B cells in the presence of T cells and monocytes. In purified B cells, IL-4 plus anti-CD40 mAb induced IgE production which was not inhibited by VIP or SOM. However, VIP or SOM, but not SP, inhibited IgE production in B cells cultured with both T cells and monocytes. Finally, the mechanism of modulation of IgE and IgG4 production was dependent on IL-4-induced switching, since neuropeptides modulated IgG4 and IgE production in surface IgG4-negative (sIgG4-) and sIgE- B cells, respectively. In contrast, modulation of IgG2 and IgG3 production was not due to switching, since neuropeptides did not affect either IgG2 or IgG3 production in sIgG2- or sIgG3- B cells, respectively.

Enhancement of in vitro spontaneous IgE production by topical steroids in patients with atopic dermatitis

BACKGROUND Atopic dermatitis (AD) is an inflammatory skin disease. Although topical steroids are widely used for AD, management of severe AD is not satisfactory because of relapse or occasional aggravation of symptoms. Moreover, glucocorticoids induce in vitro IgE production. On the other hand, topical sodium cromoglycate (SCG) solution is a safe and effective treatment for AD. METHODS We treated 43 patients with AD with SCG solution (n = 21) or with topical steroids, beclomethasone dipropionate (BD) ointment (n = 22). After 2 weeks, clinical evaluation and spontaneous immunoglobulin production by peripheral blood B cells or surface IgE+ B cells from patients in the SCG and BD groups were assessed. RESULTS Both SCG and BD treatment remarkably improved eczema. However, although SCG treatment decreased spontaneous IgE production by B cells without affecting production of IgG, IgM, or IgA, BD treatment selectively increased spontaneous IgE production. SCG treatment also decreased IgE production by surface IgE+ B cells, whereas BD treatment increased it. CONCLUSION Topical steroid treatment increases in vitro spontaneous IgE production by B cells. This indicated that topical steroids may decrease inflammation; however, a large-scale study on the effect of topical steroids on IgE production in vitro and in vivo may be necessary.

Effect of recombinant human erythropoietin on human IgE production in vitro

Recombinant human erythropoietin enhanced spontaneous IgE production 200–300% enhance ment) in cultures of peripheral blood mononuclear cells (MNC) from atopic patients. In contrast, IgG and IgA production were only slightly enhanced (30–50% enhancement), and IgM production was not affected by erythropoietin. The enhancement of IgE production by erythropoietin was indirect since it required T cells and monocytes. However, erythropoietin effect was specific since enhancement was blocked by anti‐erythropoietin antibody but not by control antibody. Interleukin‐4 (IL‐4) also enhanced spontaneous IgE production from atopic MNC. However, the enhancing effect by erythropoietin is different from that by IL‐4, since the erythropoietin effect was not blocked by anti‐IL‐4 antibody, and conversely IL‐4 effect was not blocked by anti‐erythropoietin antibody. In contrast to the enhancing effect on atopic MNC, erythropoietin failed to induce IgE production in cultures of MNC from normal donors while IL‐4 induced IgE production from normal MNC. However, when normal MNC were pre‐incubated with IL‐4. erythropoietin enhanced IgE production from IL‐4‐pre‐incubated MNC. Moreover. B cells separated from IL‐4‐pre‐incubated MNC produced IgE which was enhanced by erythropoietin. However, this effect required T cells and monocytes. These results indicate that erythropoietin could regulate ongoing IgE production in vitro by T cell‐ and monocyte‐dependent mechanisms.

Erythropoietin enhances immunoglobulin production and proliferation by human plasma cells in a serum-free medium.

The effect of recombinant human erythropoietin (Epo) on plasma cells was studied in a serum-free medium, COSMEDIUM-001 (Cosmedium). Epo enhanced both Ig production and thymidine uptake by human plasma cell lines, AF-10 and IM-9. Interleukin-6 (IL-6) enhanced both Ig production and thymidine uptake by AF-10 and IM-9, while other cytokines, including IL-1 beta, IL-2, IL-3, IL-4, IL-5, granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon-alpha (IFN-alpha) or IFN-gamma, failed to do so. However, the Epo effect was specific since Epo-induced enhancement of Ig production and thymidine uptake was blocked by the anti-Epo antibody but not by the anti-IL-6 antibody or the control antibody. Conversely, IL-6-induced enhancement was blocked by the anti-IL-6 antibody but not by the anti-Epo antibody. Epo also enhanced Ig production (IgG, IgM, and IgA) and thymidine uptake by PCA-1+ plasma cells generated in vitro. This enhancement was also blocked by the anti-Epo antibody but not by the anti-IL-6 antibody. Taken together, these results suggest that Epo enhances plasma cell responses by a different mechanism than does IL-6.

Eosinophil cationic protein inhibits immunoglobulin production and proliferation in vitro in human plasma cells

The effect of eosinophil cationic protein (ECP) on immunoglobulin (Ig) production by and proliferation of human plasma cells was studied. ECP inhibited Ig production by and proliferation of the human plasma cell lines, IM-9 and AF-10, in a dose-dependent fashion. As little as 0.05 ng/ml ECP was found to be inhibitory, and the maximal inhibition was achieved at doses of 0.1-0.5 ng/ml ECP. This inhibition was not due to cytotoxicity, since viability was always greater than 98%. Kinetic experiments demonstrated that inhibition was observable after 24 hr of culture with ECP and that the inhibitory effect of ECP was reversible. The inhibitory effect of ECP could be blocked by anti-ECP serum, but not by control serum. Of the various cytokines tested, including interleukin (IL)-1 beta, IL-2, IL-3, IL-4, IL-5, IL-6, interferon (IFN)-alpha, IFN-gamma, granulocyte-macrophage colony-stimulating factor (GM-CSF) and erythropoietin (Epo), IL-6 reversed the inhibition, while other cytokines failed to do so. ECP also inhibited Ig (IgG1, IgG2, IgG3, IgG4, IgM, and IgA) production by and proliferation of PCA-1+ plasma cells generated in vitro with a similar dose-response pattern. This inhibition also was blocked by anti-ECP serum but not by control serum, and was restored by IL-6. These results suggest that ECP may interact with IL-6 in controlling plasma cell responses.

Nerve growth factor inhibits immunoglobulin production by but not proliferation of human plasma cell lines

The effects of nerve growth factor (NGF) on human plasma cells were studied. NGF inhibited immunoglobulin (Ig) production but not thymidine uptake by human plasma cell lines IM-9 and AF-10 in a dose-dependent fashion. This NGF-induced inhibition of Ig production was specific, since inhibition was blocked by anti-NGF serum but not by control serum. Interleukin (IL)-6 did not affect Ig production by IM-9 and AF-10; however, IL-6 restored NGF-induced inhibition of Ig production. NGF also inhibited Ig production (IgG, IgM, and IgA) without affecting thymidine uptake by PCA-1+ plasma cells generated in vitro. This inhibition was also blocked by anti-NGF serum but not by control serum and was restored by IL-6. These results suggest that NGF may interact with IL-6 in control of Ig production by plasma cells.

Parathyroid hormone inhibits immunoglobulin production without affecting cell growth in human B cells

The effect of parathyroid hormone (PTH) on immunoglobulin (Ig) production and proliferation in the human B-cell lines CBL, SKW, and CESS was studied. PTH inhibited Ig production from all the B-cell lines in a dose-dependent manner during 5 days of culture. As little as 0.1 ng/ml was inhibitory. PTH also inhibited Ig production from cell lines stimulated by vasoactive intestinal peptide (VIP), interleukin 2 (IL-2), and IL-6. This inhibition was not due to decreased cell growth since proliferation was not affected and cell viability was always greater than 98%. In contrast to PTH, inactivated PTH or triiodothyronine failed to affect Ig production. Inhibition by PTH was blocked by anti-PTH serum, but not by control serum. Of the various cytokines tested, IL-4 reduced the PTH-induced inhibition of Ig production, whereas other cytokines, including IL-1 beta, IL-3, IL-5, interferon alpha (IFN-alpha), IFN-gamma, and granulocyte-macrophage colony-stimulating factor (GM-CSF), failed to do so. The reducing effect of IL-4 was blocked by anti-IL-4 antibody but not by control antibody. Moreover, IFN-alpha and IFN-gamma, but not GM-CSF, overcame the reducing effect of IL-4. PTH also inhibited IgG, IgM, and IgA production by tonsillar B cells stimulated with Staphylococcus aureus Cowan strain I (SAC) and IL-6 without affecting proliferation. This inhibition was blocked by anti-IL-4 antibody but not by control antibody. These results indicate that, in addition to its regulatory effect on calcium metabolism, PTH also acts as an immunoregulatory factor, and that it interacts with the cytokine, IL-4.

Inhibition of ongoing immunoglobulin production by eosinophil cationic protein

The effect of eosinophil cationic protein (ECP) upon ongoing immunoglobulin (Ig) production and proliferation in human B cells was studied. ECP inhibited Ig production by the human lymphoblastoid cell lines, CBL and GM-1056, in a dose-dependent fashion. In contrast, proliferation was not affected. This ECP-induced inhibition of Ig production was specific, since inhibition was blocked by anti-ECP serum but not by control serum. Interleukin (IL)-4 did not affect Ig production by CBL or GM-1056 cells; however, IL-4 reversed ECP-induced inhibition of Ig production and this reverse was blocked by anti-IL-4 antibody but not by control antibody. In contrast, other cytokines, including IL-1 beta, IL-2, IL-3, IL-5, IL-6, interferon (IFN)-alpha, and IFN-gamma, failed to reverse inhibition. ECP also inhibited spontaneous Ig production (IgM, IgG1, IgG2, IgG3, IgG4, and IgA) by tonsillar large activated B cells without affecting proliferation. This inhibition was also blocked by anti-ECP serum but not by control serum and was reversed by IL-4 specifically. These results indicate that ECP may play an important role in B cell responses.