The effects of IL-4 and IL-5 on the IgA response by murine Peyer's patch B cell subpopulations

IL-5 has been shown to specifically enhance IgA secretion in LPS-stimulated splenic B cell cultures. Maximum enhancement of IgA in such cultures, however, requires IL-4 in addition to IL-5. Because the Peyer's patches (PP), compared with spleen and lymph nodes, are enriched for precursors of IgA-secreting cells, we tested whether IL-4 and IL-5 would have a more profound effect on IgA secretion by polyclonally stimulated PP cells than spleen cells. The combination of IL-4 and IL-5 causes a comparable enhancement of IgA secretion in both LPS-stimulated PP and splenic B cell cultures. The majority of IgA secreted in LPS-stimulated PP cell cultures is derived from the sIgA- population. Furthermore, the binding high level of peanut agglutinin, germinal center subpopulation of PP cells is essentially nonresponsive to LPS, even in the presence of lymphokines; the majority of secreted IgA in these cultures is derived from the binding low level of peanut agglutinin population. In contrast to LPS-stimulated cultures, PP B cells secrete considerably more IgA than splenic B cells when polyclonally stimulated by a clone of autoreactive T cells in the presence of IL-4 and IL-5. The majority of IgA made by T cell-stimulated PP cell cultures is derived from the sIgA+ population. In these cultures, sIgA- PP cells and spleen cells secrete comparable levels of IgA and other non-IgM isotypes suggesting that sIgA- PP B cells are similar to splenic B cells in their potential to switch to IgA. In T cell-stimulated cultures the majority of IgA as well as of all other isotypes is also derived from the nongerminal center, binding low level of peanut agglutinin population.     

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.     

Dietary fructooligosaccharides induce immunoregulation of intestinal IgA secretion by murine Peyer's patch cells

Probiotic supplements induce immunological responses in the host, and dietary fructooligosaccharides (FOS) stimulate the growth of selected intestinal microflora. In this study we investigated the immunological influences of orally administrated FOS. BALB/c mice were orally administered 0-7.5% FOS for 6 weeks, and the intestinal mucosal immune responses were measured. In the 2.5%-FOS group, fecal IgA was significantly increased. IgA secretion by Peyer's patch (PP) cells was upregulated in a dose-dependent way in response to FOS and CD4+ T cells from PP showed a dose-dependent increase in production of interferon-gamma and interleukin (IL) 10, and a high response in production of IL-5 and IL-6. In contrast, FOS suppressed serum IgG1. Our findings suggest that FOS supplementation changes the intestinal environment of microflora, and leads to upregulation of IgA secretion in CD4+ PP cells in intestinal mucosa, and to suppression of the systemic immune response to type 2 helper T (Th2) dominant.     

CD11b+ Peyer's patch dendritic cells secrete IL-6 and induce IgA secretion from naive B cells

Peyer's patch (PP) dendritic cells (DCs) have been shown to exhibit a distinct capacity to induce cytokine secretion from CD4(+) T cells compared with DCs in other lymphoid organs such as the spleen (SP). In this study, we investigated whether PP DCs are functionally different from DCs in the SP in their ability to induce Ab production from B cells. Compared with SP DCs, freshly isolated PP DCs induced higher levels of IgA secretion from naive B cells in DC-T cell-B cell coculture system in vitro. The IgA production induced by PP DCs was attenuated by neutralization of IL-6. In addition, the induction of IgA secretion by SP DCs, but not PP DCs, was further enhanced by the addition of exogenous IL-6. Finally, we demonstrated that only PP CD11b(+) DC subset secreted higher levels of IL-6 compared with other DC subsets in the PP and all SP DC populations, and that PP CD11b(+) DC induced naive B cells to produce higher levels of IgA compared with SP CD11b(+) DC. These results suggest a unique role of PP CD11b(+) DCs in enhancing IgA production from B cells via secretion of IL-6.     

Selective adherence of IgA to murine Peyer's patch M cells: evidence for a novel IgA receptor

M cells represent the primary route by which mucosal Ags are transported across the intestinal epithelium and delivered to underlying gut-associated lymphoid tissues. In rodents and rabbits, Peyer's patch M cells selectively bind and endocytose secretory IgA (SIgA) Abs. Neither the nature of the M cell IgR nor the domains of SIgA involved in this interaction are known. Using a mouse ligated ileal loop assay, we found that monoclonal IgA Abs with or without secretory component, but not IgG or IgM Abs, bound to the apical surfaces of Peyer's patch M cells, indicating that the receptor is specific for the IgA isotype. Human serum IgA and colostral SIgA also bound to mouse M cells. The asialoglycoprotein receptor or other lectin-like receptors were not detected on the apical surfaces of M cells. We used recombinant human IgA1 and human IgA2 Abs and domain swapped IgA/IgG chimeras to determine that both domains Calpha1 and Calpha2 are required for IgA adherence to mouse Peyer's patch M cells. This distinguishes the M cell IgA receptor from CD89 (FcalphaI), which binds domains Calpha2-Calpha3. Finally, we observed by immunofluorescence microscopy that some M cells in the human ileum are coated with IgA. Together these data suggest that mouse, and possibly human, M cells express an IgA-specific receptor on their apical surfaces that mediates the transepithelial transport of SIgA from the intestinal lumen to underlying gut-associated organized lymphoid tissues.     

Differential regulation of IgA production by TGF-beta and IL-5: TGF-beta induces surface IgA-positive cells bearing IL-5 receptor, whereas IL-5 promotes their survival and maturation into IgA-secreting cells.

Transforming growth factor beta (TGF-beta) and IL-5 have been shown to augment IgA production by LPS-stimulated murine B cells. We investigated the effect of TGF-beta on the expression of surface Ig-isotype and IL-5 receptor on LPS-stimulated B cells. TGF-beta increased the proportion of both surface IgA-positive (sIgA+) B cells and sIgG2b+ B cells and enhanced IgA and IgG2b production by LPS-stimulated B cells. TGF-beta synergized with IL-5 only for IgA production of the seven Ig-isotypes and in combination with IL-5 caused a significant increase in the proportion of sIgA+ B cells up to 17.4%. In contrast, IL-5 decreased the proportion of sIgG2b+ B cells and sIgG3+ B cells and inhibited the production of IgG2b and IgG3 by LPS-stimulated B cells. About 50% of sIgA+ cells induced by TGF-beta expressed IL-5 receptor. They secreted peak levels of IgA and seemed to maintain long viability in the presence of IL-5; whereas TGF-beta had the opposite effects on sIgA+ B cells and down-regulated the IL-5 receptor expression. These results indicate that TGF-beta increases the number of sIgA(+)- and IL-5 receptor-positive B cells which respond to IL-5 giving rise to IgA-secreting cells and also support the notions that TGF-beta preferentially induces switching to sIgA+ B cells and IL-5 induces the maturation of postswitch sIgA+ B cells into IgA-secreting cells in a stepwise fashion.     

IL-4-dependent IgE switch in membrane IgA-positive human B cells

IgE responses by human B cells, separated according to membrane Ig classes, were analyzed in a clonal assay using EL-4 thymoma cells as helper cells, T cell supernatant, and rIL-4. In cultures seeded by means of the autoclone apparatus of the FACS, IgE responses were generated frequently by either IgM (mu+/gamma-alpha-) or IgA (alpha +/mu-)-positive B cells (16 and 14% of the Ig producing wells, respectively), but rarely by IgG (gamma +/mu-)-positive B cells (1.3% of Ig producing wells). The total amounts of Ig secreted by IgM-, IgG-, or IgA-positive cells and the total proportions of responding autoclone wells (23-27%) were comparable. All IgE secretion was IL-4 dependent. When the Ig secretion patterns from alpha +/mu- vs alpha +/mu-epsilon- B cells were compared, most autoclone wells from both types of cells produced IgA only, and similar proportions of IgA producing wells (6.2 and 6.0%) also secreted IgE. In addition, IgE restricted responses occurred 6 times more frequently with alpha +/mu- than with alpha +/mu-epsilon- cells, which suggests that membrane IgA+E double-positive, IgE committed B cells occur in vivo. The isotype pattern generated by alpha +/mu-epsilon- B cells cannot be explained by a chance assortment of separate IgA and IgE precursors or by cytophilic antibody. Thus, IL-4 dependent switch to IgE occurred frequently in IgM- or IgA-positive, but rarely among total IgG-positive, B cells. This could be relevant to IgE production in mucosal tissues rich in IgA expressing B cells.     

Distribution of substance P receptors on murine spleen and Peyer's patch T and B cells

Previous studies have demonstrated that the sensory neuropeptide substance P (SP) can modulate immune responses in vitro. Work from this laboratory has shown that SP enhances immunoglobulin synthesis by murine splenic and Peyer's patch lymphocytes stimulated with concanavalin A. One mechanism underlying these effects is the binding of SP to specific receptors on lymphocytes. We examined the distribution of SP receptors on murine T and B lymphocytes and their subsets by one and two color fluorescence-activated cell sorter analysis. The specificity and nature of binding was examined using radiolabeled SP, and competitive inhibition experiments were performed with cold SP. In cytofluorimetry experiments, both T and B lymphocytes from Peyer's patches and spleen were bound to SP, with those from Peyer's patches having a higher proportion than lymphocytes from the spleen. The majority of T cells from both organs bound SP with binding being evenly distributed between Lyt-1+ and Lyt-2+ cells. Similarly, the majority of B lymphocytes from spleen and Peyer's patches showed SP binding. There were no significant isotype-specific differences within any organ. Studies using 125I-labeled SP showed specific binding to all lymphocyte subpopulations examined. SP receptors were fewer in number on cells isolated from spleen than on cells from Peyer's patches although the dissociation constants were similar for all populations examined. These studies demonstrated that SP receptors are present both on murine T and B lymphocytes from Peyer's patches and spleen. There is no evidence for differential SP receptor expression on distinct lymphocyte subsets in spleen or Peyer's patches.     

Simultaneous induction of antigen-specific IgA helper T cells and IgG suppressor T cells in the murine Peyer's patch after protein feeding

The effects of feeding the dietary protein antigen, ovalbumin (OVA), on OVA-specific IgG and IgA immune responses involving Peyer's patches (PP) and mesenteric lymph nodes (MLN) were examined. Mice were administered soluble OVA by gastric intubation. One to 3 days later, PP, MLN, or spleen cells from these donor mice were adoptively transferred into normal syngeneic recipients. After two subsequent immunizations, spleens from the recipient mice were assayed for IgA and IgG anti-OVA plaque-forming cell (PFC) responses. None of the tissues from normal (unfed) mice had the inherent ability to alter recipients' IgG or IgA PFC responses. Within 1 day of OVA feeding, however, cells were generated in the PP that could augment recipients' IgA anti-OVA PFC responses and suppress IgG PFC responses. Three days after OVA feeding, these cells were present in MLN as well, and whereas the IgG suppressor cell also appeared to migrate to spleen, the IgA helper cell did not. The cells mediating antigen-specific IgG suppression and IgA help were both T cells but could be distinguished by surface phenotype. We therefore conclude that protein feeding induces differential, isotype-specific immunoregulation in gut-associated lymphoid tissues, part of which is mediated by an antigen-specific IgA helper T cell.     

Targeting of secretory IgA to Peyer's patch dendritic and T cells after transport by intestinal M cells

In addition to being instrumental to the protection of mucosal epithelia, secretory IgA (SIgA) adheres to and is transported by intestinal Peyer's patch (PP) M cells. The possible functional reason for this transport is unknown. We have thus examined in mice the outcome of SIgA delivered from the intestinal lumen to the cells present in the underlying organized mucosa-associated lymphoreticular tissue. We show selective association of SIgA with dendritic cells and CD4(+) T and B lymphocytes recovered from PP in vitro. In vivo, exogenously delivered SIgA is able to enter into multiple PP lining the intestine. In PP, SIgA associates with and is internalized by dendritic cells in the subepithelial dome region, whereas the interaction with CD4(+) T cells is limited to surface binding. Interaction between cells and SIgA is mediated by the IgA moiety and occurs for polymeric and monomeric molecular forms. Thus, although immune exclusion represents the main function of SIgA, transport of the Ab by M cells might promote Ag sampling under neutralizing conditions essential to the homeostasis of mucosal surfaces.     

IL-5-induced IgA synthesis by LPS-stimulated mouse B cells is prevented by protein kinase C inhibitors.

We have previously demonstrated that activation of protein kinase C (PKC) by phorbol esters induces selectively IgA synthesis by mouse B cells. In this study, we investigated the effects of a number of protein kinase inhibitors on IgA secretion induced by a recombinant murine IL-5 in LPS-stimulated mouse B cells. The results show that PKC inhibitors, such as sphingosine (SPH), staurosporine (STP) and H-7, blocked IL-5-induced IgA synthesis; the protein kinase A inhibitor HA-1004 and the inhibitor of calcium/calmodulin dependent protein kinase W-7 had no effect on IgA secretion induced by IL-5. The proliferation of the IL-5 sensitive B13 cell line in response to IL-5 was also inhibited by addition of SPH or STP or H-7. The data suggest an involvement of the PKC pathway in IL-5-induced B cell differentiation into IgA secreting cells.     

Transforming growth factor-beta 1 and IL-4 regulate the adhesiveness of Peyer's patch high endothelial venule cells for lymphocytes.

The adhesion of lymphocytes to endothelial cells lining the postcapillary high endothelial venules (HEV) is the first step in their emigration from the bloodstream into lymph nodes and Peyer's patches (PP). We have recently shown that the adhesiveness of cultured rat lymph node and PP HEV cells for thoracic duct lymphocytes can be increased significantly by pretreatment with TNF-alpha, IFN-gamma, and IL-4. In the present study we investigated the role of transforming growth factor-beta 1 (TGF-beta) on the adhesiveness of nonstimulated and cytokine-stimulated PP HEV cells for rat lymphocytes. The results indicated that at picomolar concentrations, TGF-beta significantly (p less than 0.001) decreased the ability of PP HEV cells to adhere 51Cr-labeled rat lymphocytes. Maximal inhibition was observed with a TGF-beta dose of 0.5 ng/ml and an incubation time of 6 to 12 h. TGF-beta did not affect the morphology of HEV cells and had no adverse effect on their viability. Moreover, the decrease in HEV adhesiveness by TGF-beta was reversible, with lymphocyte binding returning to control level 24 h after removal of the cytokine. The specificity of TGF-beta was confirmed by the ability of neutralizing anti-TGF-beta 1 antibody, but not control serum, to abolish the inhibitory properties of the cytokine. In addition, TGF-beta completely abrogated the increased adhesiveness of PP HEV cells normally induced by TNF-alpha or IFN-gamma. In contrast, TGF-beta had no effect on the stimulating effects of IL-4. Moreover, preincubation of PP HEV cells with TGF-beta did not alter the ability of these cells to respond to IL-4. Importantly, the adhesion of rat lymphocytes to IL-4-stimulated PP HEV cells can be blocked by pretreatment of lymphocytes with the PP-homing receptor-specific 1B.2.6 antibody whereas pretreatment of human mononuclear cells with anti-very late activation antigen-4 alpha antibody inhibited only partially the binding of these cells to the IL-4-stimulated PP HEV monolayers. Taken together, these findings strongly suggest that TGF-beta and IL-4 play important regulatory roles in lymphocyte-HEV adhesion and that the stimulatory effect of IL-4 is mediated at least in part through the increased expression of organ-specific ligands on HEV cells.     

The role of IL-5 in IgA B cell differentiation

IL-5 enhances secretion of IgA by B cells. The stage of B cell differentiation at which IL-5 enhances IgA secretion and the mechanism by which it exerts this effect are unknown. We examined these issues by separating Peyer's patch (PP) B cells into membrane IgA (mIgA)-positive and mIgA-negative cells with panning or cell sorting. When LPS was used to activate these cells, mIgA-positive PP B cells were induced by IL-5 (either as crude T cell supernatant or rIL-5 to secrete large amounts of IgA. In contrast mIgA-negative PP B cells showed no significant amount of IgA secretion with IL-5. In addition, rIL-5 did not cause expression of mIgA by mIgM-bearing B cells. The mechanism involved in enhancement of IgA secretion was evaluated by utilizing an ELISPOT assay to quantitate IgA secreting cells. Both unsorted PP B cells and mIgA-positive PP B cells, when incubated with IL-5, showed an increase in the number of IgA-secreting cells that was proportional to the increase in total secreted IgA. However, LPS-activated PP mIgA-positive B cells, when incubated with rIL-5, showed no increase in proliferation, as measured by [3H]thymidine incorporation indicating that the increase in IgA-secreting cells after incubation with IL-5 occurred not as a result of proliferation but rather through promotion of terminal differentiation. Thus, IL-5 acts as a differentiation factor on B cells which have already undergone isotype switch to IgA B cells, promoting differentiation into IgA-secreting cells with resultant increased IgA secretion.     

Regulation of mucosal IgA production in vitro by autoreactive immunoregulatory T cells from murine Peyer's patches

In this study, we investigated whether Peyer's patch-derived T-cell subsets participate in vitro in self major histocompatibility (MHC) class II antigen (Ag)-mediated immunoregulatory circuits for gut-mucosal IgA isotype selection in the presence of Peyer's patch (PP)-derived syngeneic surface immunoglobulin M (sIgM)-bearing B cells. When fresh (in vitro unstimulated) sIgM-bearing B cells were cocultured with fresh, PP-derived L3T4+ Vicia villosa-nonadherent (VV-) T cells (T helper (Th) cells), the production of all class-specific immunoglobulins (Ig), but, in particular, IgA, was enhanced two- to sixfold. This augmented Ig production was, however, reduced by nearly 50% when fresh PP-derived Lyt2+ VV-T cells (suppressor T cells) were added. Furthermore, addition of PP-derived L3T4+ VV+ and Lyt 2+ VV+ T cells abrogated, by nearly 100%, the suppression induced by the Lyt 2+ VV-T cells (contrasuppression). When lipopolysaccharide (LPS)-stimulated, PP-derived sIgM-bearing B cells were cocultured with the Th cells, the production of each class-specific Ig was similarly enhanced, but Ig levels exceeded those obtained with cultures of the unstimulated B cells (P less than 0.001). Anti-I-A or anti-I-E monoclonal antibody (mAb) inhibited the induction of each immunoregulatory T-cell effector activity (P less than 0.001), and anti-I-A/E inhibited it synergistically. Thus, unstimulated fresh PP-derived T cells appear to be activated and then to exert T-cell effector functions in the sequential development of helper, suppressor, and contrasuppressor immunoregulatory networks in the presence of PP-derived sIgM B cells and, particularly, LPS-preactivated sIgM B cells. Based on the blocking effect of anti-I-A and/or anti-I-E mAb on the induction of each T-cell-mediated immunoregulatory function in class-specific Ig production, it appears that the autoreactive (self MHC class II Ag-reactive) activation of PP T cells evoked by Ia Ag on PP sIgM B cells largely controls mucosal IgA production by the latter cells. Furthermore, this immunoregulation by autoreactive effector T cells, especially the L3T4+ VV- helper T cell, may play a significant role in vivo in gut-mucosal IgA isotype production.     

Estrogen induces normal murine CD5+ B cells to produce autoantibodies

Females have better humoral immune responses and are more susceptible to autoimmune diseases than males. Normal female mice (C57BL/6J, C3H/HeJ, and NZW) have significantly increased spontaneous autoimmune plaque-forming cells (APFC) to mouse erythrocytes pretreated with bromelain (Br-ME) in spleen, peritoneal exudate cell, and bone marrow compared to their male counterparts. A minor subpopulation of B cells, CD5+ B, is thought to produce this autoantibody. As determined by dual color flow cytometry, increased APFC to Br-ME in females is not due to quantitative increase of CD5+ B cells. Rather, it is due to increased numbers or percentages) of CD5+ B cells producing these autoantibodies, because CD5+ B cells from females produced greater numbers of APFC to Br-ME than equal numbers of cells derived from males. The increased autoantibody production in females can be attributed to the effect of estrogen on the immune response because this hormone markedly augments APFC to Br-ME in intact or orchidectomized males. Male hormone had little effect. Importantly, estrogen did not increase the numbers of B or CD5+ B cells but augmented the ability of B cells to produce this response. This was verified when a T cell-depleted B cell fraction or fluorescence-activated cell sorter purified CD5+ B cells from estrogen-treated mice proved more efficient in the production of APFC to Br-ME. These results suggest that the number of CD5+ B cells committed to produce autoantibodies to Br-ME is increased under the influence of estrogen. This is the first demonstration that estrogen can augment the production of natural autoantibodies in normal mice. The overall augmented humoral immune responses in females and the B cell hyperactivity in female predominant autoimmune diseases appears to be due to estrogen.     

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.     

Increased frequency of surface IgA-positive plasma cells in the intestinal lamina propria and decreased IgA excretion in hyper IgA (HIGA) mice, a murine model of IgA nephropathy with hyperserum IgA

Because abnormalities of mucosal immunity have been suggested in human IgA nephropathy, we examined the involvement of mucosal immunity in IgA deposition to the kidney in hyper IgA (HIGA) mice, which was established as a mouse model for human IgA nephropathy with hyperserum IgA. The number of surface IgA+B220- lymphocytes in the intestinal lamina propria (LP) of HIGA mice increased 2.7-fold at 30 wk of age as compared with those at 10 wk of age, whereas normal mice did not show such increase. The surface IgA+B220- LP lymphocytes spontaneously secreted IgA in culture. Morphological studies showed that the surface IgA+B220- lymphocytes of murine intestinal LP are identical with plasma cells (PCs). About 20% of IgA+B220- PC in LP expressed both Mac-1 and CD19, suggesting that they may derive from peritoneal B-1 cells. Cell cycle study on intestinal IgA-PCs using bromodeoxyuridine revealed no difference between HIGA mice and normal mice, suggesting that the high frequency of IgA-producing PCs in HIGA mice is not due to enhanced proliferation or prolonged survival of IgA-producing PCs in LP. In addition, IgA secretion into the gut lumen of HIGA mice decreased drastically (to one forth) with aging. These data suggest that the increased number of intestinal IgA-producing PCs and the down-regulation of IgA excretion into the intestinal lumen might synergistically contribute to the hyperserum IgA in HIGA mice and resultant IgA deposition to the kidney.     

Differential glycosylation of two glycoproteins synthesized by murine B cells in response to IL-4 plus IL-5

We sought to determine whether selected cytokines, known to stimulate profoundly B-cell activation and differentiation, also have as yet unrecognized effects upon the glycosylation of secreted Ig and/or membrane-associated proteins. The glycosylation of both secreted IgM and membrane-bound MHC Class-I synthesized by CH12LX cells was detected by enzyme-lectin conjugates in immunoabsorption assays. Stimulation of B cells with IL-4 plus IL-5 significantly decreases the terminal glycosylation of secreted IgM, whereas LPS has a minor effect, despite the fact that both stimuli are equipotent for IgM secretion. Neither LPS nor IL-4 plus IL-5 affect MHC Class-I expression. However, IL-4 plus IL-5 substantially increases the terminal glycosylation of MHC Class-I produced from both mIgM(+)and mIgA(+)CH12LX cells. LPS has no or a modest effect on the terminal glycosylation of MHC Class-I produced from CH12LX cells. These results suggest that Th(2)-derived cytokines differentially influence the glycosylation of secreted and membrane-associated glycoproteins of B cells. In turn, this might elucidate the basis of aberrant glycosylation reported in conditions such as IgA nephropathy, cancer and rheumatoid arthritis.