For 10 weeks a 25-year-old man had been suffering from tiredness, fatigue, nausea and a 16 kg weight loss. Erythrocyte sedimentation rate (83/133 mm), serum C-reactive protein (5.5 mg/dl) and creatinine (5.05 mg/dl) were all elevated. He also had proteinuria (1120 mg daily), sterile leukocytosis and a creatinine clearance of 10 ml/min. Renal biopsy showed interstitial nephritis and bone marrow biopsy revealed non-caseous epithelioid-cell granulomas. 14 days after admission he developed acute iritis in the right eye. Other causes having been excluded, the diagnosis of tubulo-intestinal nephritis with uveitis (TINU syndrome) was made. The clinical symptoms and laboratory findings improved within a few days of the start of glucocorticoid treatment (initially, 100 mg prednisone daily, reduced to 5 mg within 30 days). The patient was discharged after 8 days in good general condition.

An 18-year-old boy, who had severe abdominal pain for 18 months associated with marked weight loss, was found to have a stenosed ileal sling on double-contrast radiology of the small intestine. At operation a plate-like tumour was extending from the pancreas to the aortic bifurcation. Histological examination of removed tissue revealed retractile mesenteritis (mesenteric panniculitis; liposclerotic mesenteritis). The symptoms regressed and the patient again gained weight under immunotherapy with 1 mg/kg of prednisone and 2 mg/kg of cyclophosphamide daily. Subsequently, under prednisone alone, there was a recurrence, which responded within five weeks to daily 60 mg prednisone and 125 mg cyclophosphamide. The patient remains symptom-free on 125 mg cyclophosphamide and 10 mg prednisone daily.

HLA-B27 molecules expressed on the T2 mutant cell line do not have peptides. Such empty HLA-B27 molecules were not recognized by an HLA-B27-restricted cytotoxic T-lymphocyte (CTL) clone (auto-1) derived from synovial fluid. To test for peptide dependency of the clone, B27-T2 cells were incubated with a panel of 48 different peptides. This lack of stringency was compared with that of a peptide-dependent monoclonal antibody, B27.M2. Positive B27.M2 reactivity resulted when the B27-T2 cells were incubated with two peptides: RRKAMFEDI and RRMGPPVGHR, derived from Chlamydia HSP60 and human ribonucleoprotein, respectively. Because of the limited availability of CTL versus monoclonal antibody, the specificity of B27.M2 was studied in greater detail. The importance of the HLA-B27 heavy chain in antibody recognition of class I-peptide complexes was demonstrated by site-directed mutagenesis. The stringency of the peptide residues was tested by making analogs of each of the nine residues in RRKAMFEDI, creating a panel of 180 analogs. Although stringency was highest for the sixth position, as many as six different amino acids provided positive reactivity. These results indicate that immune recognition of HLA-B27-peptide complexes might have rather low stringency for the peptide sequences. In theory, then, pathogen-derived peptides which induce autoimmunity by generating autoreactive CTL might not share much sequence similarity with the responsible self peptides.

We report the case of a 41-year old patient who underwent orthotopic liver transplantation because of decompensated liver cirrhosis due to chronic HCV-infection. Severe acute allograft rejection was insufficiently controlled by cyclosporine A, steroids and a 6-day regimen of OKT 3 monoclonal antibody therapy. As a consequence immunosuppressive therapy was switched to FK 506 in a dose of 3 mg bid. The FK 506 concentration in whole blood consistently ranged between 5.1 and 7.8 ng/ml. Seven weeks after the onset of FK 506 therapy the patient developed severe diabetes mellitus with fasting blood glucose levels up to 640 mmol/l. The C-peptide was elevated reflecting a higher than normal insulin secretion. Intravenous insulin therapy with application of up to 85 units regular insulin per day was initiated. Because of the severe diabetes immunosuppression was changed back to cyclosporine A. After six weeks the patient did no longer require insulin and showed an entirely normal glucose tolerance test, C-peptide and Hb A1-level. This case shows that the diabetogenic side effect of FK 506 is more pronounced than that of cyclosporine A. We propose to change immunosuppressive therapy to cyclosporine A in cases of FK 506 induced severe diabetes mellitus, since long-term prognosis of many transplant recipients may depend on side effects of the immunosuppressive agents.

OBJECTIVE In reactive arthritis (ReA), synovial fluid-derived bacteria-specific CD4+ and CD8+ T cells have been studied intensively in recent years. We have addressed the question whether gamma delta-TCR+ lymphocytes could contribute to antibacterial or anti-self cytotoxicity in the affected joints of patients, with spondylarthropathies. METHODS T cell clones were derived by random cloning from the synovial fluids of one patient with Yersinia-induced ReA, one patient with a Yersinia-induced flare up of pre-existing ankylosing spondylitis, and one patient with ankylosing spondylitis. Eight clones with a CD3+, alpha beta-TCR-, CD4-, CD8- and gamma delta-TCR+ phenotype (all expressing V gamma 9) were tested in a standard 52Cr-release assay using autologous or allogeneic B cell lines, CIR-B27, Daudi cells, and RJ.225 cells. RESULTS Four gamma delta-TCR+ clones killed both autologous and allogeneic target cells when infected with live Yersinia or Salmonella and also uninfected Daudi cells expressing GroEL heatshock protein. One clone was specific for Yersinia-infected targets. Three gamma delta-TCR+ clones were cytotoxic when uninfected autologous or allogeneic targets were employed. Polymorphic "classical" MHC class I or class II molecules were not used as restriction elements. CONCLUSION We conclude that, upon in vivo contact with bacteria such as Yersinia and Salmonella, synovial gamma delta-T lymphocytes are activated and contribute to antibacterial immunity via specific target cell lysis. Furthermore, anti-self cytolytic gamma delta-T cells could participate in the clearance of stressed and detrimental cells in the arthritic joint or, alternatively, could support the chronicity of autoimmune arthritis.

In the differential diagnosis of infection-related arthritis (infectious arthritis, viral arthritis, reactive arthritis or Reiter's syndrome, Lyme disease) various laboratory methods are applied for the detection of the inciting antigen, specific antibodies or microbespecific T-lymphocytes. In infectious (septic) bacterial or fungal arthritis, the definitive diagnosis can be made only by recovering the organism from the synovial fluid or membrane. Also, in reactive arthritis following extraarticular infection with Yersinia, Salmonella, Shigella, Campylobacter, or Chlamydia, one of the major shifts in perception of disease pathogenesis has been the detection of bacterial determinants by immunological methods and polymerase chain reaction (PCR) actually within the joint. In sexually acquired reactive arthritis, the etiologic diagnosis should be based on the direct detection of the pathogen (mainly C. trachomatis) from the urogenital smear specimen. For clinical routine, serological tests for bacteria specific antibodies (IgM and IgA class) are often necessary to show recent or persistent infection with the triggering pathogen. However, a cautionary note regarding the diagnostic significance of antibacterial antibody profiles has been sounded in several studies because of the high prevalence of bacteria-specific antibodies in the healthy population. The same problem may arise in the interpretation of virus-specific antibodies in the differential diagnosis of acute polyarthritis. Antigen-specific proliferation of synovial fluid lymphocytes can confirm the clinical diagnosis in patients with reactive arthritis and Lyme disease, although unspecific proliferation to several bacteria can also be observed in reactive arthritis as well as in many other arthritis.

OBJECTIVE:To study the frequency of Klebsiella pneumoniae-responsive T cells in the peripheral blood (PB) of ankylosing spondylitis (AS) patients compared with that in healthy HLA-B27+ donors, and to examine T lymphocyte clones (TLC) derived from AS patient synovial fluid (SF) for the presence of Klebsiella reactivity.METHODS:Limiting dilution analysis of PB T cells in 8 patients with active AS and in 8 HLA-B27+ healthy subjects was used to determine the frequency of PB T cells responsive to K pneumoniae and Escherichia coli GroEL. SF T cells from a patient with active AS were cloned, and 125 TLC were characterized in proliferation assays.RESULTS:There were fewer T cells in the PB of AS patients that reacted with K pneumoniae than in the PB of healthy HLA-B27+ subjects. The frequencies of E coli GroEL-responsive T cells were approximately 5-10 times lower in all subjects tested (healthy donors and AS patients), but without significant differences between the 2 groups. Two CD4+ TLC that recognized K pneumoniae (1 cross-reactive with E coli) as well as 3 TLC that recognized GroEL (2 CD4+, 1 T cell receptor gamma/delta+) were isolated from the SF of a patient with actige AS.CONCLUSION:Our results indicate that there is a quantitative reduction of K pneumoniae-responsive T cells in the PB of AS patients as compared with healthy controls. This may reflect a defective peripheral T cell defense in the immune response to Klebsiella and may allow bacterial antigens to reach the synovium, where they initiate specific T cell responses.

In the differential diagnosis of infection-related arthritis (infectious arthritis, viral arthritis, reactive arthritis or Reiter's syndrome, Lyme disease) various laboratory methods are applied for the detection of the inciting antigen, specific antibodies or microbe-specific T-lymphocytes. In infectious (septic) bacterial or fungal arthritis, the definitive diagnosis can be made only by recovering the organism from the synovial fluid or membrane. Also, in reactive arthritis following extraarticular infection with Yersinia, Salmonella, Shigella, Campylobacter, or Chlamydia, one of the major shifts in perception of disease pathogenesis has been the detection of bacterial determinants by immunological methods and polymerase chain reaction (PCR) actually within the joint. In sexually acquired reactive arthritis, the etiologic diagnosis should be based on the direct detection of the pathogen (mainly C. trachomatis) from the urogenital smear specimen. For clinical routine, serological tests for bacteria specific antibodies (IgM and IgA class) are often necessary to show recent or persistent infection with the triggering pathogen. However, a cautionary note regarding the diagnostic significance of antibacterial antibody profiles has been sounded in several studies because of the high prevalence of bacteria-specific antibodies in the healthy population. The same problem may arise in the interpretation of virus-specific antibodies in the differential diagnosis of acute polyarthritis. Antigen-specific proliferation of synovial fluid lymphocytes can confirm the clinical diagnosis in patients with reactive arthritis and Lyme disease, although unspecific proliferation to several bacteria can also be observed in reactive arthritis as well as in many other arthritis.

We present the case of a female patient with a diagnosis of systemic lupus erythematosus (SLE) at the age of 54 years. At the age of 63 years, she suffered from malignant thymoma and 3 years after removal of the thymoma a diagnosis of pure red cell aplasia (PRCA) was established. This is, to our knowledge, the first report of the occurrence of SLE, thymoma and PRCA in the same patient. The case is discussed with regard to the already known associations between these diseases.

For 10 weeks a 25-year-old man had been suffering from tiredness, fatigue, nausea and a 16 kg weight loss. Erythrocyte sedimentation rate (83/133 mm), serum C-reactive protein (5.5 mg/dl) and creatinine (5.05 mg/dl) were all elevated. He also had proteinuria (1120 mg daily), sterile leukocytosis and a creatinine clearance of 10 ml/min. Renal biopsy showed interstitial nephritis and bone marrow biopsy revealed non-caseous epithelioid-cell granulomas. 14 days after admission he developed acute iritis in the right eye. Other causes having been excluded, the diagnosis of tubulo-intestinal nephritis with uveitis (TINU syndrome) was made. The clinical symptoms and laboratory findings improved within a few days of the start of glucocorticoid treatment (initially, 100 mg prednisone daily, reduced to 5 mg within 30 days). The patient was discharged after 8 days in good general condition.

SUMMARY Hyporesponsiveness to a universe of bacterial and dietary antigens from the gut lumen is a hallmark of the intestinal immune system. Since hyperresponsiveness against these antigens might be associated with inflammation, we studied the immune response to the indigenous intestinal microflora in peripheral blood, inflamed and non-inflamed human intestine. Lamina propria monocuclear cells (LPMC) isolated from inflamed intestine but not peripheral blood mononuclear cells (PBMC) of IBD patients with active inflammatory disease strongly proliferated after co-culture with sonicates of bacteria from autologous intestine (BsA), Proliferation was inhibitable by anti-MHC class II MoAb, suggesting that it was driven by antigen, LPMC from adjacent non-inflamed intestinal areas of the same IBD patients and PBMC or LPMC isolated from non-inflamed intestine of controls and patients with IBD in remission, in contrast, did not proliferate, PBMC or LPMC which had been tolerant to bacteria from autologous intestine, however, strongly proliferated after co-culture with bacterial sonicates from heterologous intestine (BsH). This proliferation was associated with an expansion of CD8+ T cells, increased expression of activation markers on both CD4+ and CD8+ lymphocyte subsets, and production of IL-12, interferon-gamma (IFN-γ), and IL-10 protein. These results show that tolerance selectively exists to intestinal flora from autologous but not heterologous intestine, and that tolerance is broken in intestinal inflammation. This may be an important mechanism for the perpetuation of chronic IBD.

Although the pathogenesis of psoriasis vulgaris is still unknown, several characteristics point to an immunologically mediated process. Epidermal psoriatic lesions are characterized by a hyperproliferation of keratinocytes and an infiltration of T lymphocytes and granulocytes. Because the former may be mediated in part by lymphokines secreted by T cells, we have focused our interest on the in vivo and in vitro cytokine secretion patterns of T lymphocytes from psoriatic lesions. In five patients T lymphocytes were obtained from epidermal specimens. The cells were propagated with lectin and irradiated feeder cells and subsequently cloned by limiting dilution. The resulting T-cell clones were phenotypically and functionally characterized. Our data show that the majority of T-cell clones were CD4+ (74%), whereas only 25% were CD8+ and 1% were CD4-/CD8-. Also, we have further investigated the cytokine secretion pattern of T-cell lines or CD4+ T-cell clones, respectively. All cells tested produced interferon-gamma whereas only a minority secreted interleukin (IL)-4. Moreover, these cells produced high amounts of IL-2 but only little or no IL-10 or tumor necrosis factor-alpha. To correlate these data with the in vivo situation, biopsies from psoriatic lesions of five patients were investigated for the presence of the mRNA of IL-4, IL-10, and interferon-gamma using the polymerase chain reaction. In these biopsies only the mRNA for the Th1 cytokine interferon-gamma but not for the Th2 cytokines IL-4 and IL-10 could be detected. Identical experiments were performed to test the in vivo cytokine production of synovial fluid mononuclear cells of two patients with arthropathia psoriatica. Again, only the mRNA for interferon-gamma but not IL-4 could be detected. This indicates that T cells involved in psoriasis exhibit a Th1-like cytokine secretion profile.

The term seronegative spondylarthropathies is used for an entity of rheumatic syndromes of peripheral joints and the spine (ankylosing spondylitis, reactive arthritis, Reiter's syndrome, arthritis in psoriasis and in inflammatory bowel disease) which are strongly associated with the MHC class I molecule HLA-B27. However, the mechanisms whereby HLA-B27 confers disease susceptibility have so far remained unknown. There is strong evidence that gut inflammation and infection with gram-negative bacteria play a role in the induction of B27-associated disease. HLA-B27, like other MHC class I molecules, physiologically binds antigenic peptides in its binding groove and presents them to CD8+ T lymphocytes. Consequently, if the disease association with HLA-B27 arises from its role as a T-cell restriction element, synovial fluid CD8+ rather than CD4+ T cells should play a prominent pathogenetic role and should be detectable within the affected joints. In this paper, recent studies on bacteria-specific cytotoxic T cells and on peptide binding to HLA-B27 are reviewed. Particular emphasis is laid on the role of HLA-B27 restricted synovial CD8+ T cells with specificity for bacterial antigens or autoantigens. These cytotoxic T cells could provide a missing link in the pathogenesis of the spondylarthropathies and could now serve as tools to identify the critical antigenic epitopes of bacterial and self peptides which are involved in disease induction.

Reactive arthritis is a usually self-limited sterile inflammation of joints that follows certain bacterial gastrointestinal or urogenital infections. The immunopathogenesis involves CD4+ T cells, which mediate an antigen-specific TH1 response to bacterial constituents within the joint. Properties of the arthritogenic bacteria and the physicochemical characteristics of the bacterial antigens may contribute to the development of reactive arthritis.

Gamma/delta TCR bearing T lymphocytes represent a T-cell subset whose functional relevance remains unclear. Nevertheless these T cells may play a role in the early immune response against bacteria. Until now the regulatory mechanisms on this response have not been investigated. The study described here evaluated the immunoregulatory effects of Interleukin-10 on gamma/delta and alpha/beta TCR-positive T-cell clones and freshly isolated peripheral-blood mononuclear cells (PBMC). IL-10 has been shown previously to inhibit lectin and antigen-induced proliferation and cytokine production by alpha/beta T cells. The results outlined below show that rhIL-10 strongly inhibits lectin-induced production of IFN-gamma, TNF-alpha, IL-2, and to a lesser degree proliferation and IL-4 production of both T-cell subsets. As IL-10 did not inhibit proliferation but at the same time strongly suppressed cytokine production in various experiments, the hypothesis that it could function as a growth factor for human T cells as has been described for murine thymocytes was tested. The data demonstrate that, although the gamma/delta T-cell clones tested do not produce IL-10 they can use it as a growth factor in combination with IL-2, IL-4 or alone. Furthermore, IL-10 has the same properties on human alpha/beta T-cell clones and PBMC. In summary, it is shown that IL-10 has pleiotropic effects on gamma/delta and alpha/beta TCR+ T cells by inhibiting lectin-induced cytokine production and by acting as a growth factor for these cells alone or in combination with IL-2 or IL-4.

The arthritis-predisposing HLA-B27 consists of a heavy chain, a small peptide, and the monomorphic beta 2-microglobulin (beta 2-m). CTLs and a mAb, Ye-2, which recognize the complex with specificities both for the heavy chain and for the peptide, are available. The beta 2-m is in noncovalent association with the heavy chain at multiple points and is exchangeable with free beta 2-m outside of the complex. The purpose of our experiments was to test whether mutant beta 2-m capable of modulating HLA-B27 activity could be created. Eighteen recombinant mutants of the human beta 2-m were experimentally generated. In 14 of these, mutations were at or near residues that are either contact residues or interface residues with the heavy chain. Relative to the parent beta 2-m, two-thirds of the mutants showed reduced ability to exchange into HLA-B27 complexes. However, at least four of them induced more than 80% decrease in Ye-2 Ab reactivity. Two mutants were able to induce a minor decrease in susceptibility to lysis by four CTL clones. One of the CTL clones was autoreactive. Two of the CTL clones were specific for HLA-B27 cells experimentally infected with arthritis-causing Yersinia enterocolitica. These results indicate that certain beta 2-m residues play an indirect role in peptide presentation, although they are not directly associated with the peptide residues.