Anti-CD2 and anti-CD3 induced T cell cytotoxicity of human intraepithelial and lamina propria lymphocytes.

The effector function of immunocompetent cells in the gut mucosa has not yet been defined. The cytotoxic function of these cells might be important in the normal immune response and could be relevant to the mucosal damage seen in inflammatory conditions. The cytotoxic function of isolated intraepithelial and lamina propria mononuclear cells in six and 18 hour assays after the addition of various stimuli that interact with the human leukocyte antigens CD2 and CD3 on the mucosal effector cells was investigated. T cell phenotypes were determined using CD4, CD8, and HML1 to characterise cells of the appropriate compartments. Anti-CD3 and phytohaemagglutinin can induce toxic activity of lamina propria lymphocytes in most individuals after six hours and in all individuals after 18 hours. Anti-CD2, anti-CD3, and phytohaemagglutinin are similarly effective at triggering lamina propria lymphocytes. Intraepithelial lymphocytes contain predominantly CD8 and HML1 positive T cells, differentiating phenotypically intraepithelial lymphocytes from lamina propria lymphocytes. Intraepithelial lymphocytes are not cytotoxic at six hours, but have a toxic function comparable with lamina propria lymphocytes after 18 hours with all three triggers. Intraepithelial lymphocytes from inflamed mucosa (Crohn's disease and diverticulitis) mediate significantly reduced cytotoxicity in vitro compared with normal mucosa, whereas lamina propria lymphocyte toxicity is not different. Reduced numbers of cytotoxic cells and reduced reactivity to the trigger substances used after in vivo activation or cold target inhibition could explain the observed differences between intraepithelial lymphocytes from inflamed and uninflamed mucosa. Changes in cell mediated cytotoxicity of intraepithelial lymphocytes and lamina propria lymphocytes may be involved in the mucosal damage in these inflammatory conditions.

Immunoglobulin G (IgG), IgG1, and IgG2 determinations from endoscopic biopsy specimens in control, Crohn's disease, and ulcerative colitis subjects.

Acute exacerbations of chronic inflammatory bowel disease (ulcerative colitis and Crohn's disease) are characterised by an increase in immunoglobulin G (IgG) positive cells in the mucosa, whereas uninflamed mucosa of inflammatory bowel disease patients displays only moderately increased or normal numbers of these cells. Previous data suggest that acute exacerbations of ulcerative colitis and Crohn's disease can be distinguished by different IgG subclass expression of mucosal immunocytes and a different IgG subclass production pattern of lamina propria lymphocytes. A procedure to obtain enough intestinal mononuclear cells from biopsy specimens to measure in vitro IgG and IgG1 production in control subjects and various patient groups has been established. IgG2 could be measured in Crohn's disease and ulcerative colitis only, as the concentrations in control subjects were below the sensitivity of the ELISA method. We found that IgG and IgG1 production correlated with the degree of local inflammation in both diseases, even in slightly inflamed mucosa, compared with control subjects. The proportion of IgG1 subclass was significantly increased in severely inflamed mucosa of both ulcerative colitis and Crohn's disease patients. A major difference between Crohn's disease and ulcerative colitis mucosa is apparent in mild or no inflammation. In Crohn's disease mucosa in remission, the IgG1/IgG ratio is comparable with that in controls, yet ulcerative colitis mucosa still displays significantly increased proportions of IgG1. In addition, the IgG2/IgG ratio is 0.12 in ulcerative colitis and 0.19 in Crohn's disease patients. The results show the dependence of local IgG and IgG1 production on the degree of inflammation and that an increase in subclass IgG1 in ulcerative colitis is present at all stages, including remission. These findings support the hypothesis that different immunoregulatory mechanisms are involved in Crohn's disease and ulcerative colitis. Environmental stimuli or genetic background may be responsible for the observed differences.

Cefodizime stimulates subpopulations of cells mediating spontaneous or antibody-dependent cytotoxicity in patients with bacterial infections.

The influence of cefodizime (CDZ) on spontaneous cell-mediated cytotoxicity (SCMC) and antibody-dependent cell-mediated cytotoxicity (ADCC) was investigated in nine patients with conditions predisposing to infection (T-cell deficiency, humoral immune deficiency, myeloproliferative syndrome, kidney or liver damage, or chronic pulmonary disease). SCMC, using the K562 and LIK cell lines as targets, and ADCC, using the LIK cell line, were measured at baseline and after ten days of therapy with CDZ for lower respiratory tract infections. Six patients were cured; clinical outcome was not evaluable in the other three. SCMC lysis of K562 cells did not change significantly. SCMC and ADCC lysis of LIK (lymphoblastoid) cells both increased significantly. CDZ selectively stimulated a subpopulation of NK cells in this population of immunocompromised patients with lower respiratory tract infections.

[Treatment of severe Raynaud syndrome in scleroderma or thromboangiitis obliterans with prostacyclin (prostaglandin I2)]. / Behandlung des schweren Raynaud-Syndroms bei Sklerodermie oder Thromboangiitis obliterans mit Prostacyclin (Prostaglandin I2).

Eleven patients with severe Raynaud's syndrome were treated with intravenous infusion of prostacyclin (Prostaglandin I2). Raynaud's syndrome was caused by inflammatory diseases such as progressive systemic sclerosis (N = 9) or thromboangiitis obliterans (N = 2). Five patients had acral ulcerations. Treatment with prostacyclin lead to immediate cessation of acral pain in all patients if doses of 5-6 ng/kg/min were tolerated. In 7 out of 11 patients there was a long-term analgesic effect with clinical improvement of Raynaud's syndrome. In three of five patients we achieved healing of the ulcerations within a few weeks. Plasmaconcentrations of prostaglandin F1-alpha, the main metabolite of prostacyclin, were about 10 times above normal during infusion and returned to normal levels within 30 min after the end of the infusion, in spite of the prolonged clinical effect. Therefore, prostacyclin alone cannot be responsible for the long-term clinical benefit. (Parts of this publication were published as an abstract and presented at the 23rd Congress of the Deutsche Gesellschaft für Rheumatologie (15).

[The small intestine as an immune organ]. / Der Dünndarm als Immunorgan.

The immune system of the gastrointestinal tract (gut-associated lymphoid tissue--GALT) differs from the peripheral immune system in a number of points, and can also react largely independently of the latter. The lymphatic cells of GALT are both strictly compartmentalized (Peyer's patches, lymphatic follicles), and diffusely distributed within the mucosa. The organized lymphatic tissue represents the afferent component, the diffuse lymphatic tissue the efferent component, of the intestinal immune response. A marked recirculation behavior (homing) of the intestinal lymphocytes makes it clear hat GALT is simply part of a more comprehensive common mucosal immune system, the mucosa-associated lymphatic tissue (MALT). At the center of a description of the pathophysiology of the small bowel as an immune organ is Crohn's disease. By way of example, current immunological aspects of immunoregulation, specific and unspecific cytotoxic, that is, tissue-injurious, immune reactions, as well as immunoglobulin isotype and immunoglobulin G subclass differentiation are discussed.

Alterations in serum immunoglobulin G subclasses in patients with ulcerative colitis and Crohn's disease.

We have examined the concentration of immunoglobulin G (IgG) subclass antibodies in the sera of 27 patients with ulcerative colitis and 21 patients with Crohn's disease as well as in 11 normal controls and 11 patients with systemic lupus erythematosus. In comparison with a control mean serum IgG1 concentration of 5173 micrograms/ml, patients with ulcerative colitis exhibited a significantly increased mean serum concentration of 7924 micrograms/ml (p less than 0.05), whereas patients with Crohn's disease had a near normal mean serum IgG1 level of 5898 micrograms/ml. In contrast, control sera had a mean IgG2 level of 2477 micrograms/ml and ulcerative colitis sera had a similar IgG2 level of 2269 micrograms/ml, whereas Crohn's disease sera had a significantly increased mean IgG2 level of 5111 micrograms/ml (p less than 0.05). Patients with systemic lupus erythematosus, like those with ulcerative colitis, had a markedly elevated serum IgG1 level of 15,594 micrograms/ml (p less than 0.001) without a significantly increased IgG2 serum level (3271 micrograms/ml). Neither ulcerative colitis nor Crohn's disease sera exhibited altered levels of IgG3 or IgG4. These data show that alterations in IgG subclass concentrations occur in the sera of patients with active, untreated inflammatory bowel disease, similar to the previously noted changes in the IgG subclasses secreted by lymphocytes from involved inflammatory bowel disease intestinal specimens.

Ulcerative colitis specific cytotoxic IgG-autoantibodies against colonic epithelial cancer cells.

Serum antibodies cytotoxic to the colon cancer cell line RPMI 4788 were studied in 42 patients with ulcerative colitis, 61 patients with Crohn's disease, 27 patients with other inflammatory diseases (disease-controls) and 22 healthy controls. Cytotoxicity of antibodies towards RPMI 4788 was studied by means of a chromium release assay using peripheral blood mononuclear leucocytes of healthy subjects as effector cells. Using a four hour antibody dependent cell mediated cytotoxicity assay sera from 29% of ulcerative colitis patients contained antibodies cytotoxic for the target, while only 3% of the Crohn's patients and 6% of the disease controls and non of the healthy controls were positive. When an 18 hour assay was applied, however, not only 40% of ulcerative colitis patients, but also 14% of Crohn's patients and 21% of disease controls were found positive. The reactive antibody in the four hour assay was mainly of the IgG class, which points at a classical antibody dependent cell mediated cytotoxicity mechanism. In the 18 hour cytotoxic assay IgG and particularly IgM antibodies were found to be reactive. This suggests that in the latter case other cellular cytotoxic mechanism might be involved. There was a significant inverse correlation between the appearance of the ulcerative colitis restricted IgG-anticolon epithelial cell antibodies (four hour assay) and the disease activity (p less than 0.01). Absorption studies showed that the reactive antigen is not specific for ulcerative colitis colonic tissue, but is similarly found in Crohn's bowel tissue, and to a lower extent in normal bowel, liver and kidney. The reactive antigen, however, could not be detected in brain and lymphoblastoid cells.

[Non-steroidal anti-inflammatory agents increase intestinal permeability]. / Nicht-steroidale Antiphlogistika erhöhen die Darmpermeabilität.

51Cr-EDTA activity was measured in urine and blood of patients receiving non-steroidal anti-inflammatory (NSA) drug treatment and of healthy subjects and other patients (controls), after oral intake of 51Cr-EDTA, for the purpose of deciding whether NSA treatment increases urinary excretion of oral 51Cr-EDTA as an expression of increased intestinal permeability. 51Cr-EDTA activity in urine and blood of patients with rheumatoid arthritis (13) being treated with NSA was significantly higher (similar to results in 13 patients with Crohn's disease) than that of a control group (14) of patients with rheumatoid arthritis without such treatment (9) and patients without rheumatic disease (5). Both in patients with rheumatoid arthritis receiving NSA drugs and patients with Crohn's disease there was a highly significant correlation between urinary and blood activity. There was no effect of NSA drugs on renal function. The results indicate that NSA drugs increase interenterocytic permeability to an extent comparable to permeability abnormalities in Crohn's disease.

[Intestinal spontaneous cell-mediated cytotoxicity and its immunoregulation in patients with chronic inflammatory bowel disease and controls]. / Untersuchungen der intestinalen spontanen zellvermittelten Zytotoxizität und ihrer Immunregulation bei Patienten mit chronisch-entzündlichen Darmerkrankungen und Kontrollen.

Isolated intestinal mononuclear cells (IMNL) exhibit spontaneous cell-mediated cytotoxicity (SCMC) in one third of the cases. IMNL suppress the SCMC of peripheral blood lymphocytes. The suppression by IMNL of patients with chronic inflammatory bowel disease is diminished.

Immune status in Crohn's disease. VI. Immunoregulation evaluated by multiple, distinct T-suppressor cell assays of lymphocyte proliferation, and by enumeration of immunoregulatory T-lymphocyte subsets.

In 28 patients with Crohn's disease, 6 patients with ulcerative colitis, and 34 healthy controls, immunoregulatory function of peripheral blood mononuclear cells was investigated by evaluating the suppression of lymphocyte proliferative responses to mitogens (phytohemagglutinin, concanavalin A, pokeweed mitogen) and to allogeneic lymphocytes (mixed lymphocyte culture) using simultaneously five functional assays as follows: (a) spontaneous T-suppressor-cell activity, (b) concanavalin A-generated T-suppressor-cell activity, both with (3000 rads) and without irradiation of suppressor cells, and (c) allogeneic mixed lymphocyte culture-generated T-suppressor-cell activity, again both with and without irradiation. Concanavalin A- and mixed lymphocyte culture-generated T-suppressor-cell activities were evaluated both in the autologous and the allogeneic system. In addition, using monoclonal antibodies, we determined proportions of T-helper and T-suppressor/cytotoxic lymphocytes. Inactive patients did not differ from normal either in the proportions of immunoregulatory lymphocytes or in the suppression of the various lymphocyte proliferative responses in any of the five T-suppressor-cell assays evaluated. In patients with active disease, however, an impairment of suppression of phytohemagglutinin-, pokeweed mitogen-, and mixed lymphocyte culture-stimulated proliferation of autologous lymphocytes was observed in the concanavalin A-generated, irradiated suppressor assay. In the spontaneous suppressor assay, suppression of phytohemagglutinin- and concanavalin A-stimulated lymphocyte proliferation was significantly lower in active disease than in remission. Thus, in peripheral blood of patients with Crohn's disease who are in remission, there is no indication for an immunoregulatory defect in any of the evaluated assay systems. Single selective, moderate defects in suppression of proliferation of various lymphocyte subpopulations are restricted to active disease.

[Immunologic aspects of chronic inflammatory intestinal diseases]. / Immunologische Aspekte bei chronisch entzündlichen Darmerkrankungen.

Aetiology as well as pathogenesis of Crohn's disease and ulcerative colitis are unclear as yet. Recent studies suggest that a transmissible factor present in Crohn's disease lymph nodes produces lymphoma and immune complex glomerulonephritis in nu/nu mice. Furthermore, sera of those patients contain an antibody, that recognizes an "antigen(s)" in the murine lymphoma and glomerular immune complexes. In ulcerative colitis a disease-specific colonic tissue bound antibody directed against specific antigens of colonic mucosa from patients with ulcerative colitis but not from patients with Crohn's disease and normal colon has been found. Contrary to earlier reports up to date knowledge gives no indication for a primary immunological defect of functions of the cell mediated immunity or unspecific immune system, although impairment of some immunological parameters appears due to the disease, both in Crohn's disease and in ulcerative colitis. This holds true for peripheral blood mononuclear cells. Results obtained with mononuclear cells isolated from the gut are highly controversial and at present do not allow final conclusions.

[Serodiagnosis of Crohn's disease]. / Untersuchungen zur Frage einer Serodiagnostik des Morbus Crohn.

Three serological tests, recommended as being of diagnostic value for Crohn's disease, were evaluated in 39 patients with Crohn's disease and--as controls--in 27 patients with ulcerative colitis, 45 healthy persons and 65 patients with inflammatory diseases other than Crohn's disease or ulcerative colitis. The tests were the determination of (1) serum antibodies to pseudomonas-like organisms (PLO) by means of indirect immunofluorescence; (2) agglutinating serum antibodies to 4 strains of anaerobic gram-positive coccoid rods (species of Eubacterium, Peptostreptococcus and Coprococcus); and (3) serum antibodies to perinuclear antigens in buccal mucosa of Crohn's disease patients by immunofluorescence. The results indicate that the occurrence of high-titer antibodies to PLO is reasonably sensitive for Crohn's disease, but has a low specificity, and that antibodies to perinuclear antigens in buccal mucosa have both low sensitivity and specificity. However, the occurrence of agglutinins to 4 strains of anaerobic grampositive coccoid rods is significantly higher in Crohn's disease than in ulcerative colitis, patients with other diseases and healthy controls. Thus the determination of these agglutinins does not discriminate between Crohn's disease and ulcerative colitis; but it is a serodiagnostic adjunct in the diagnosis of chronic inflammatory bowel diseases.

Selected bacterial antibodies in Crohn's disease and ulcerative colitis.

Agglutinins to four strains of anaerobic gram-positive coccoid rods (species of Eubacterium, Peptostreptococcus and Coprococcus) were found in significantly higher frequency in Crohn's disease (CD) than in ulcerative colitis (UC) and in other diseased control subjects and were virtually absent in apparently healthy subjects. When the posterior probability of having CD was calculated on the basis of these agglutination reactions, 64% of patients with CD and 34% of patients with UC but only 10% of diseased controls and none of the healthy controls were regarded as 'probable' or 'definite' cases of CD. However, the posterior probability of CD did not sharply differentiate between CD and UC but indicated chronic inflammatory bowel disease. Factors contributing to the appearance of these agglutinins in CD were also evaluated. The findings would indicate the importance of a damaged intestinal mucosal barrier for the production of these agglutinins, provided the antigens are present in the intestine. No significant differences were observed between the occurrence of antibodies to pseudomonas-like organisms (PLO) in CD and the various control groups. The study could not add further evidence to the hypothesis of a possible aetiopathogenic role of PLO in CD.

In vitro inhibition of protein catabolism by alpha 2-macroglobulin in plasma from a patient with posttraumatic acute renal failure.

Rapid proteolytic digestion of three subunits of phosphorylase kinase was shown in plasma ultrafiltrates from a patient who was admitted with multiple traumatic injuries, respiratory insufficiency, hemorrhagic shock and acute renal failure. The observed cleavage of phosphorylase kinase may be a consequence of protease-antiprotease imbalance. After an initial determination of 80 mg/dl the alpha 2-macroglobulin values were too low to be detected. Addition of purified alpha 2-macroglobulin to the ultrafiltrates resulted in complete inhibition of phosphorylase kinase digestion in vitro. Aprotinin did not inhibit proteolytic digestion. Daily dialysis did not ameliorate the observed protein catabolism of this patient. These findings may have clinical application in hypercatabolic states, if alpha 2-macroglobulin becomes available in a form suitable for human use.