Spontaneous secretion of IgG subclasses by intestinal mononuclear cells: differences between ulcerative colitis, Crohn's disease, and controls

Spontaneous IgG and IgG subclass secretion patterns by isolated intestinal mononuclear cells (MNC) from control and inflammatory bowel disease (IBD) specimens were examined. Intestinal MNC from IBD specimens spontaneously secreted more total IgG than did control intestinal MNC. This increased spontaneous IgG secretion by ulcerative colitis intestinal MNC was primarily due to markedly increased production of IgG1. Slightly increased secretion of IgG3, but not IgG2 by ulcerative colitis intestinal MNC was present when compared with control and Crohn's disease intestinal MNC. In contrast, Crohn's disease intestinal MNC exhibited increased spontaneous secretion of all the IgG subclasses examined, with IgG2 being predominant.     

Antibody secretion by human intestinal mononuclear cells from normal controls and inflammatory bowel disease patients

Inflammatory bowel disease (IBD) intestinal mononuclear cells (MNC) exhibit decreased spontaneous IgA secretion with an increased percentage of monomeric IgA and IgA subclass 1 in both ulcerative colitis and Crohn's disease patients. When compared with control intestinal MNC, a marked increase in spontaneous secretion of IgG is observed from IBD MNC. The greatest increase in spontaneous IgG secretion is seen with ulcerative colitis intestinal MNC, due to the secretion of large amounts of IgG subclass 1. Crohn's disease intestinal MNC have increased IgG subclass 2 secretion. Similar differences in IgG subclass concentrations also occur in the sera of active, untreated, IBD patients. Therefore, major alterations occur with regard to spontaneous antibody secretion of IgA and IgG subclasses in IBD. Because intestinal MNC comprise a unique immunologic compartment, it will be important to better understand the regulatory mechanisms, effector capabilities, and inducing antigens involved in intestinal IgA and IgG subclass secretion in IBD.     

Immunoglobulin production by isolated intestinal mononuclear cells from patients with ulcerative colitis and Crohn''s disease.

We studied immunoglobulin production by isolated intestinal mononuclear cells from 25 patients with active inflammatory bowel disease (IBD) and 17 controls undergoing surgical resections for intestinal tumour or other disorders. Normal ileal intestinal mononuclear cells spontaneously produced greater amounts of IgA and IgM than did normal colon cells. In cells from patients with IBD there was a significantly reduced IgA production, but production of IgG was enhanced in both colon and ileum. The alteration in IgA and IgG production in IBD was confirmed by comparing the immunoglobulin production by mononuclear cells from inflamed with that from non-inflamed areas of mucosa in six patients with distal ulcerative colitis. The proportion of IgA-containing cells in isolated intestinal mononuclear cells from IBD mucosa was less than normal. However, the proportion of IgG-containing cells from IBD mucosa was not increased in isolated intestinal mononuclear cells although they produced more IgG than normal mucosal cells. Our study showed an altered pattern of immunoglobulin production by intestinal mononuclear cells isolated from patients with inflammatory bowel disease.     

Immunology and Immunopathology of the Intestines: Antibody Secretion by Human Intestinal Mononuclear Cells from Normal Controls and Inflammatory Bowel Disease Patients

Inflammatory bowel disease (IBD) intestinal mononuclear cells (MNC) exhibit decreased spontaneous IgA secretion with an increased percentage of monomeric IgA and IgA subclass 1 in both ulcerative colitis and Crohn's disease patients. When compared with control intestinal MNC, a marked increase in spontaneous secretion of IgG is observed from IBD MNC. The greatest increase in spontaneous IgG secretion is seen with ulcerative colitis intestinal MNC, due to the secretion of large amounts of IgG subclass 1. Crohn's disease intestinal MNC have increased IgG subclass 2 secretion. Similar differences in IgG subclass concentrations also occur in the sera of active, untreated, IBD patients. Therefore, major alterations occur with regard to spontaneous antibody secretion of IgA and IgG subclasses in IBD. Because intestinal MNC comprise a unique immunologic compartment, it will be important to better understand the regulatory mechanisms, effector capabilities, and inducing antigens involved in intestinal IgA and IgG subclass secretion in IBD.     

The nature of the natural killer (NK) cell of human intestinal mucosa and mesenteric lymph node.

The relationship of the mononuclear cell (MNC) from human intestinal mucosa and mesenteric lymph node mediating anti-K-562 activity with that of peripheral blood has been assessed. Depletion of macrophages did not alter the measured cytotoxicity confirming that the effector cells were lymphocytes. Complement lysis of Leu 7 and Leu 11b coated cells reduced intestinal natural killer (NK) activity by a similar degree to that of peripheral blood but mesenteric lymph node NK activity was affected to a lesser extent. The response in NK activity of mucosal and nodal MNC to short incubation with lymphoblastoid interferon was similar to that for peripheral blood MNC. Twenty-four hours incubation of MNC with low concentrations of purified interleukin-2 (IL-2) consistently augmented intestinal and nodal NK activity but failed to augment that of peripheral blood MNC. No differences between the inhibitory effects of cAMP and prostaglandin E2 on NK activity from the three sites were seen. In addition, inhibition of cyclo-oxygenase activity with indomethacin had no effect on NK activity of intestinal and peripheral blood MNC while the lipoxygenase inhibitor, nordihydroguaiaretic acid, suppressed intestinal and peripheral blood NK activity similarly. In conclusion, anti-K-562 activity by intestinal MNC is mediated by NK cells with similar phenotypic and functional properties to those of peripheral blood. However, the increased sensitivity of mucosal NK cells to IL-2 suggests that higher proportions of NK cell precursors may be present in intestinal MNC populations.     

Characterization of antigen-presenting activity of intestinal mononuclear cells isolated from normal and inflammatory bowel disease colon and ileum.

Antigen-presenting activity in mononuclear cells, isolated from normal and inflamed human ileum and colon, has been characterized using allogeneic mixed lymphocyte reaction with resting T cells as responders. Greatest proliferation was induced by fibronectin-adherent (macrophage-enriched) cells, and least by fibronectin non-adherent (macrophage-depleted) cells and by mononuclear cells depleted of macrophages by panning with monoclonal antibody 3C10. When intestinal mononuclear cells and allogeneic T cells were incubated in large numbers, clusters were observed. These clusters contained cells with a dendritic morphology that were strongly HLA-D-positive and which also stained with macrophage-specific monoclonal antibodies 3C10, EMB11 and Y1/82A. These cells were closely associated with proliferating T cells. Studies comparing mononuclear cells isolated from normal and inflamed colonic mucosa suggest that the latter may have enhanced antigen-presenting capacity.     

Effect of recombinant human erythropoietin on human IgE production in vitro.

Recombinant human erythropoietin enhanced spontaneous IgE production (200-300% enhancement) 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.     

Increased spontaneous secretion of rheumatoid factor by intestinal lamina propria mononuclear cells from Crohn's disease but not ulcerative colitis patients.

Increased levels of rheumatoid factors (RF) have been observed in the serum of Crohn's disease but not ulcerative colitis patients, and have been proposed to relate to an increased state of intestinal lymphocyte activation. We have therefore examined the spontaneous in vitro secretion of RF by intestinal lamina propria mononuclear cells (MNC) isolated from specimens from control and inflammatory bowel disease (Crohn's disease, ulcerative colitis) patients. Normal intestinal lamina propria MNC spontaneously secrete rheumatoid factors of different isotypes during 14 days of in vitro culture (9.7 ng/ml IgA RF, 11.6 ng/ml IgM RF and 64.6 ng/ml IgA anti-Fc (IgG)). In matched studies intestinal MNC isolated from normal large bowel exhibited significantly greater levels of RF synthesis and secretion in vitro than normal small bowel intestinal MNC. A large increase in spontaneous RF secretion was observed from Crohn's disease intestinal MNC (21.4 ng/ml IgA RF, 21.4 ng/ml IgM RF, and 108.15 ng/ml IgA anti-Fc (IgG)), when compared with normal controls. The amount of RF secreted was dependent on the amount of inflammatory activity of the bowel specimens, from which the MNC were isolated (198.3 ng/ml of IgA anti-Fc(IgG) from involved versus 50.0 ng/ml from matched non-involved tissue). Ulcerative colitis MNC released decreased amounts of RF (7.1 ng/ml IgA RF, 6.2 ng/ml IgM RF, and 42.3 ng/ml IgA anti-Fc(IgG)). These observations using isolated intestinal MNC may explain the findings of RF changes in the sera of inflammatory bowel disease patients. Our observations support the hypothesis of a heightened state of activation in normal intestinal lamina propria MNC, which is further increased in active Crohn's disease. The dissimilarities observed between Crohn's disease and ulcerative colitis may indicate fundamental differences in disease pathophysiology and will lead to further studies exploring intestinal immunoregulatory properties of RF.     

Investigation of the expression of IL-1beta converting enzyme and apoptosis in normal and inflammatory bowel disease (IBD) mucosal macrophages.

Activated mucosal macrophages are derived from circulating monocytes and appear to play a major role in the pathogenesis of IBD. We have recently shown that IBD, but not normal, mucosal macrophages express the active form of IL-1beta converting enzyme (ICE) and are therefore capable of releasing mature IL-1beta. ICE expression by other mucosal cell types is unknown. Active ICE expression has also been implicated in apoptosis. The aim of this study was to investigate ICE expression (using an antibody that recognizes both active and precursor forms) in normal and IBD mucosa and to determine whether ICE-expressing macrophages are undergoing apoptosis. Normal and active IBD mucosal cells, in tissue sections and after isolation, were studied by immunohistochemistry and flow cytometry. In the mucosa, macrophages were the predominant ICE-expressing cell type. In contrast to normal, most IBD mucosal macrophages expressed ICE. Of IBD colonic macrophages 11.8 +/- 3.2%, and of normal colonic macrophages 6.6 +/- 0.6% expressed Apo2.7, a marker for apoptotic cells. Similar data were obtained when annexin V was used to identify cells undergoing apoptosis. DNA fluorescence flow cytometric analysis of normal and IBD lamina propria cells showed the presence of only small hypodiploid DNA peaks. We conclude that in the human intestinal mucosa, macrophages are the predominant ICE-expressing cell type. Expression of the active form of ICE and macrophage apoptosis are not interdependent. One mechanism of loss of resident macrophages from normal mucosa and of recruited macrophages from IBD mucosa is by apoptosis.     

Human rib bone marrow mononuclear cells spontaneously synthesize and secrete IgE in vitro.

We have examined spontaneous secretion of IgE by human rib bone marrow mononuclear cells (MNC). Bone marrow MNC from nine out of 12 rib specimens synthesized and secreted substantial amounts of IgE during 14 days of in vitro culture. The 14-day supernatants from these bone marrow MNC contained a mean of 2589 pg/ml of IgE (n = 12) with a maximum production of 15,408 pg/ml of IgE compared with small amounts of IgE (80-200 pg/ml) produced by similarly cultured normal and inflammatory bowel disease intestinal lamina propria MNC. Using two rib specimens, time-course studies revealed spontaneous secretion of IgE to be minimal during the first 2 days of culture (152 pg/ml), followed by a steady increase between days 4 (517 pg/ml) and 14 (3588 pg/ml). The addition of pokeweed mitogen resulted in 72% suppression of spontaneous IgE production by bone marrow MNC. The bone marrow MNC isolated from the ribs consisted of 22% Leu12+ (B) cells of which 3.2% were surface IgE positive. Staining for cytoplasmic immunoglobulin revealed 1% of the bone marrow MNC to be cytoplasmic IgE+. The presence of IgE-bearing and IgE-secreting MNC in human bone marrow is consistent with the observation that allergen-specific IgE-mediated hypersensitivity is adoptively transferred by human bone marrow transplantation and demonstrates the usefulness of human bone marrow MNC for examination of IgE secretory and regulatory events.     

High frequency of spontaneous interferon-gamma-producing cells in human tonsils: role of local accessory cells and soluble factors.

The frequency of mononuclear cells (MNC) spontaneously secreting interferon-gamma (IFN-gamma) has been examined in freshly isolated cell suspensions from human palatine tonsils. Two-site reverse enzyme-linked immunospot (ELISPOT) analyses, involving short term (20 h) incubation of MNC in the absence of any added exogenous stimulus, revealed that tonsillar MNC suspensions contain exceptionally large numbers of cells secreting IFN-gamma. No significant differences were observed when comparing the frequency of IFN-gamma-producing cells between cell suspensions obtained from hyperplastic and tonsillitis specimens. Cell-sorting experiments disclosed that spontaneous tonsillar IFN-gamma production was essentially contributed by CD4+ T cells, and required the presence of accessory cells and/or soluble factors to be detected. Thus, depletion of plastic adherent cells or monocytes from the tonsillar MNC suspensions resulted in reduced numbers of detectable IFN-gamma-secreting cells. Addition of very small numbers of autologous monocytes restored spontaneous IFN-gamma production in tonsillar MNC cultures depleted of monocytes. Neutralization of endogenous IL-1 beta and IL-2, as well as blocking of the IL-2 receptor, also decreased IFN-gamma production from unfractionated tonsillar cells. Addition of exogenous IL-1 beta restored IFN-gamma production in cultures of tonsillar MNC depleted of plastic adherent cells. Furthermore, IL-1 beta synergized with IL-2 by tonsillar MNC depleted of plastic adherent cells. Furthermore, IL-1 beta synergized with IL-2 by increasing intracellular as well as cell-free levels of IFN-gamma in cultures of unfractionated tonsillar MNC. This study further establishes that the tonsils are highly active immunological organs containing large numbers of T cells spontaneously producing IFN-gamma whose detection is contingent upon the presence of functional accessory cells. It also demonstrates that concomitant production of IL-1 beta and IL-2 occurs in tonsils and is necessary to maintain ongoing synthesis and extracellular accumulation of IFN-gamma in these organs.     

Regulation of intestinal immunoglobulin production in response to dietary ovalbumin.

The effects of oral administration of ovalbumin (OVA) on intestinal immunoglobulin production was examined. Balb/c mice, bred and reared on an OVA-free diet, received either one dose or 14 consecutive daily intragastric doses of 25 mg OVA/dose. Single dose administration of OVA resulted in significant suppression of total immunoglobulins in the intestinal mucosa, particularly of the IgA isotype, although a very low titre anti-OVA IgG class antibody response was induced. After multiple peroral immunisations, there was more intestinal anti-OVA antibody induction and less suppression of total immunoglobulins. However, all the anti-OVA antibody was of the IgG isotype. In vitro production of mucosal immunoglobulins was not significantly reduced over 5 days, compared with controls, in either single or multiple administration groups, suggestive of a loss of T suppressor cell function in culture. Prior adoptive transfer of splenic and lymph node cells from mice preimmunised with OVA was capable of abrogating the local suppression of immunoglobulin production in vivo. Although adoptive transfer of bovine serum albumin-sensitised cells could also overcome some of the local suppression, complete restoration of normal immunoglobulin levels was not achieved. These data suggest that single oral administration of a novel dietary antigen induces a transient, non-specific suppression of intestinal immunoglobulin production, which can be overcome by antigen-specific T cells.     

Factors affecting the spontaneous cell-mediated cytotoxicity of intestinal mononuclear cells.

This study was performed to determine what factors related to the enzymatic isolation technique and assay conditions may influence the measurement of spontaneous cell-mediated cytotoxicity (SCMC) of mononuclear cells (MNC) isolated from human intestinal mucosa. In 18 studies, the SCMC of freshly isolated cells was 1.8 +/- 0.4% but increased to 12 +/- 3% following 24 hr culture without a change in the proportion of cells with the NK phenotype (Leu-7+). The SCMC tended to plateau with more prolonged culture. Culturing peripheral blood (PB) MNC for 24 hr did not alter SCMC nor the proportion of Leu-7+ cells. However, the suppression of the SCMC of PBMNC preincubated with the supernatant of the collagenase digestion of intestinal mucosa was completely reversed by 24 hr culture. Intestinal MNC were found to suppress the SCMC of autologous PBMNC in mixing experiments. However, 24 hr culture did not affect the degree of suppression and the proportion of T cells of the suppressor-cytotoxic phenotype (Leu-2a+) was also unchanged. It is concluded that the SCMC of intestinal MNC may be accurately assessed following 24 hr culture of the cells to allow recovery from the suppressive influences of the isolation process and that this does not introduce other artefactual problems. However, suppression of cytotoxicity within the assay may result in an underestimation of the SCMC of intestinal MNC when compared to that of PBMNC.     

Subtractive screening reveals up-regulation of NADPH oxidase expression in Crohn's disease intestinal macrophages

Macrophages play a central role during the pathogenesis of inflammation. In normal intestinal mucosa surface expression of typical macrophage markers such as CD14, CD16, CD11b or T-cell co-stimulatory molecules such as CD80 or CD86 is low indicating anergy and low pro-inflammatory activity of these cells. During inflammatory bowel disease (IBD) the mucosa is invaded by a population of macrophages displaying these markers, secreting higher cytokine levels and representing an activated cell population. CD33(+) cells (macrophages) were isolated from normal and Crohn's disease mucosa and mRNA was isolated by polyT magnetic beads. A subtractive screening was performed subtracting mRNA from normal macrophages from those of Crohn's disease macrophages. Oxidative burst activity was determined by flow cytometry. Seventy clones were obtained by the subtractive mRNA screening. Sequencing showed > 99% homology to mRNA of monocyte chemoattractant protein-1 (MCP-1) for three clones. Five clones obtained by subtraction revealed > 99% homology to mRNA of cytochrome b (subunit gp91). Differential expression of the cytochrome b subunit gp91 and the cytosolic NADPH oxidase subunit p67 was confirmed by RT-PCR and 'virtual' Northern blots. The fluorescence ratio of stimulated versus unstimulated cells was 0.9 +/- 0.16 in control macrophages indicating a lack of oxidative burst activity. In Crohn's disease this ratio was significantly increased to 1.80 +/- 0.8 (P = 0.004) confirming the molecular data. In conclusion NADPH oxidase mRNA is down-regulated or absent in macrophages from normal mucosa correlating with a lack of oxidative burst activity. In IBD macrophage-oxidative burst activity is increased and NADPH oxidase mRNA induced. Inhibition of NADPH oxidase could be a new therapeutical target in IBD and reduce mucosal tissue damage in active IBD.     

Immunopathology of human inflammatory bowel disease

Conclusions Mucosal pIgA, and thereby secretory immunity, are disfavored in IBD as revealed by decreased J-chain expression and strikingly increased IgG production by the mucosal immunocytes in both UC and CD. Moreover, a significant shift from IgA2 to the less-resistant IgA1 subclass takes place. Preferential overproduction of IgG1 is seen in UC; and apical deposits of this isotype on the surface epithelium, together with activated complement, is suggestive of a cytotoxic attack related to a brushborder antigen. Comparison of identical twins, discordant with regard to clinical expression of UC, further suggests that this IgG1 response is genetically determined. The IBD lesions furthermore contain many recently recruited and activated T cells as well as monocyte-like macrophages (L1⁺CD14⁺) with elevated capacity for production of proinflammatory cytokines (IL-1 and TNF-) and ROM. Together all these changes reflect less restricted extravasation of precursor cells because of altered expression of adhesion molecules on the microvasculature. Isolated and cultured intestinal endothelial cells subjected to proinflammatory cytokines show enhanced ICAM-1 expression as well as induction of functional E-selectin and MHC class II molecules with antigen-presenting properties.The immunopathology of IBD thus involves severely altered mucosal homeostasis, apparently reflecting break of tolerance to the indigenous microbial flora, and in addition it appears to include an autoimmune component in UC. The origin of abrogated immunological tolerance at the mucosal level may be alterations both in leukocyte extravasation and in antigen-presenting mechanisms induced by aberrant expression of endothelial and epithelial MHC class II molecules, as well as changed profiles of co-stimulatory molecules on macrophages. A shift from B7.2 on mucosal resident APC to B7.1 on the recently recruited macrophages, may particularly disfavor Th2 immune responses and their associated down-regulatory cytokines. In conclusion, perturbation of a tightly controlled cytokine network with abnormal crosstalk between several mucosal cell types, is probably the first step of a progressive immunopathological development in IBD, but the initiation of this series of events remains undefined.     

Macrophage depletion decreases IgG anti-DNA in cultures from (NZB x NZW)F1 spleen cells by eliminating the main source of IL-6.

We have studied the role of macrophages in the production of IgG anti-DNA autoantibodies by (NZB x NZW)F1 mice (B/W). One of the main features of the systemic lupus erythematosus (SLE)-like disease that affects these mice, is the presence of circulating IgG autoantibodies and immune complexes, which lead to renal failure and death by the age of 8-9 months. IgG autoantibodies are produced without in vitro stimulation by total spleen cells from these mice when they reach the age of 6 months. We have demonstrated that IL-6 increases the production of IgG autoantibodies in cultures of splenic purified B cells from the old B/W mice. The aim of this study was to show the involvement of macrophages in the production of IL-6 and consequently in the production of IgG anti-DNA antibodies in vitro. We show that elimination of the macrophages by different treatments led to reduction of the content of IL-6 in the supernatants as well as of IgG anti-DNA autoantibodies. Addition of fresh, splenic or peritoneal macrophages restored the production of autoantibodies in macrophage-depleted cultures from old B/W mice. There were no differences in the capacity of IL-6 production between macrophages from old or young B/W mice, but an important difference was observed between peritoneal and splenic macrophages, where the former produced much higher levels of IL-6, and consequently were more potent inducers of IgG autoantibodies. The present results reinforce the role of macrophages and IL-6 in the production of IgG anti-DNA autoantibodies in B/W mice. The implications of these results in the pathogenesis of the disease are discussed.     

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.     

Suppression of in-vitro antibody production by pokeweed mitogen-stimulated human bone-marrow mononuclear cells. Evidence for a soluble suppressor substance

The spontaneous synthesis and secretion of immunoglobulin by human bone-marrow mononuclear cells (MNC) in vitro, as well as its suppression by the addition of pokeweed mitogen (PWM), were previously reported by this laboratory. In the present study we demonstrate that this suppression is mediated by a soluble substance elaborated by marrow MNC stimulated with PWM. Marrow MNC were pulsed for 1 hr with PWM, washed and recultured for 7 days in media without PWM. The culture supernatants were collected by centrifugation and filter sterilized before addition to fresh marrow MNC in the in vitro antibody synthesis assay. The 14-day assay culture supernatants were then subjected to a solid phase radioimmunoassay to determine the immunoglobulin content. The suppressor substance was non-specific as to immunoglobulin isotype and was not genetically restricted. Suppressor activity was diminished by heating the supernatants at 56 degrees for 1 hr. The activity could be elaborated by cells subjected to 1000 R or 2000 R before or after 1-hr incubation with PWM. While the addition of PWM anytime during the culture period would suppress IgA production at the level produced up to that time, the suppressor substance only suppressed IgA production when added during the first 4 days of culture. The addition of indomethacin had no effect on the suppressor activity indicating that the activity was not mediated by prostaglandin. Including human fibroblast interferon or hydrocortisone in the assay cultures had no effect on IgA production or its suppression by PWM. We concluded that the lectin-induced suppression was mediated by a marrow-derived suppressor substance (MDSS).     

In vitro immunoglobulin production by mononuclear cells in rheumatoid arthritis

Since patients with rheumatoid arthritis (RA) exhibit serum hypergammaglobulinemia and autoantibody (rheumatoid factor) production, we compared elaboration and control of in vitro RA mononuclear cell (MNC), Ig assayed by enzyme-linked immunoassays or by hemolytic plaque formation, in 37 RA patients and 17 normal subjects. We found (1) RA spontaneous plaque-forming cells were significantly reduced (RA 344 vs normal 627 PFC/10(6) MNC, P less than 0.002); (2) RA spontaneous IgG and IgM (but not IgA) elaboration was significantly diminished (IgG RA 339, normal 776; IgM RA 255, normal 869 ng/ml, P less than 0.001; IgA RA 87, normal 124); (3) RA stimulated IgG and IgM production (but not IgA) was also decreased (IgG RA 2434, normal 3862, P less than 0.06; IgM RA, 1676, normal 3323, P less than 0.005; IgA RA 1859, normal 2315); (4) reduced RA Ig elaboration was not clearly due to altered numbers of T or non-T cells, age, medications, clinical features of disease, or response kinetics; (5) relative improvement of RA in vitro IgG, but not usually IgM, secretion followed removal of adherent cells, addition of indomethacin or addition of mitomycin C-treated T cells; (6) MNC from synovial fluids, but not bone marrows, exhibited spontaneous Ig production in excess of stimulated synovial fluid cellular or peripheral blood Ig elaboration. These observations indicate selective impairment of peripheral blood MNC IgG and, particularly, IgM secretion in RA. This defect appears to reflect accessory cell influences which differ from normal as well as the sequestration of primed or activated cells in the synovial fluid.