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Cryoglobulinemic vasculitis (CV) is a rare immune complex disease of small vessels (capillaries, venules or arterioles) with detection of cryoglobulins (CG). These are serum proteins that precipitate at temperatures below the normal body temperature. The laboratory diagnostics are logistically challenging because the temperature of the blood sample must be maintained continuously at 37 °C until arrival in the laboratory to prevent early precipitation of the proteins with adsorption to corpuscular blood components. Cryoglobulins can be divided into three classes (types I-III), with each class associated with specific underlying diseases and symptom complexes. Cryoglobulinemia can be caused by hematological, virological or autoimmune diseases and mixed forms also occur. The most common cause to date is a hepatitis C infection. Treatment of the underlying disease is obligatory, with antiviral treatment of hepatitis C offering the rare possibility of causal treatment. Depending on the severity of cryoglobulinemia, immunosuppressive therapy is indicated to prevent permanent damage caused by the inflammation., (© 2022. The Author(s), under exclusive licence to Springer Medizin Verlag GmbH, ein Teil von Springer Nature.)

Background and Aims: Multiple clinical trials have demonstrated the efficacy and safety of tenofovir disoproxil fumarate (TDF) in chronic hepatitis B (CHB). However, long-term efficacy and safety data for TDF in real-life clinical practice are limited., Methods: Prospective German field practice study in CHB-mono-infected patients. Patients were TDF-naïve but could have been treated previously with other HBV antivirals., Results: Efficacy analysis included 400 patients; 301 (75 %) completed 36 months of TDF treatment. Both treatment-naïve and treatment-experienced patients showed a rapid decline in HBV DNA within 3 months of TDF initiation. After 36 months, HBV DNA < 69 IU/mL was achieved by 91 % of treatment-naïve patients (90 and 92 % in hepatitis B "e" antigen [HBeAg]-positive and [HBeAg]-negative, respectively) and 96 % of treatment-experienced patients (93 and 97 %, respectively). Three patients experienced virologic breakthrough, all with reported non-compliance. Overall, 5.7 % HBeAg-positive and 2.2 % HBeAg-negative patients lost hepatitis B surface antigen. Safety data were consistent with the known TDF safety profile; the most commonly reported adverse events possibly related to TDF were fatigue (2.0 %) and headache (2.0 %). Few patients (1.3 %) experienced renal-related adverse reactions. Creatinine clearance remained relatively stable over time; patients responded favorably where TDF was dose adjusted per label for decreased creatinine clearance., Conclusions: TDF showed a favorable tolerability profile and induced rapid and sustained suppression of HBV DNA in patients with CHB treated for up to 3 years in routine clinical practice, irrespective of treatment history. Efficacy and safety in this heterogeneous patient population were consistent with data from clinical trials.

Background & Aims: Despite continuous high-risk behavior, a subgroup among people who inject drugs (PWID) remains seronegative for hepatitis C virus (HCV) suggesting that a state of "natural resistance" to HCV Infection may exist. Homozygosity for KIR2DL3 and its ligand HLA-C1 group alleles has been associated with control of HCV infection, however, the mechanism mediating this protective effect remained unclear., Methods: Peripheral NK cells from PWID (n=104) were phenotypically and functionally characterized by multicolor flow cytometry. Expression levels of the NK cell receptor ligands were analysed in liver biopsies and primary human hepatocytes., Results: HCV seronegative PWID (n=34) had increased levels of KIR2DL3(+)NKG2A(-) NK cells compared to healthy controls (n=10; p<0.001) and PWID with chronic (n=38; p<0.001) or resolved infection (n=37; p<0.001). There was an inverse correlation between the frequency of KIR2DL3(+) and NKG2A(+) NK cells (r=-0.53; p<0.0001). Importantly, expression of HLA-E, the ligand for NKG2A, was significantly upregulated in liver biopsies of HCV infected patients (n=51) compared to HBV infected patients (n=22; p<0.01) and correlated with HCV viral load (r=0.32; p<0.0029). In functional analyses KIR2DL3(-)NKG2A(+) NK cells but not KIR2DL3(+)NKG2A(-) NK cells were significantly inhibited by HLA-E ligation. Accordingly, interferon gamma secretion of NK cells from PWID with chronic infection but not from HCV seronegative PWID was significantly suppressed in the presence of HLA-E., Conclusions: KIR2DL3(+)NKG2A(-) NK cells are not sensitive to HLA-E-mediated inhibition and may thereby control early HCV infection prior to seroconversion and result in an apparent state of "natural resistance" to HCV in PWID., (Copyright © 2014. Published by Elsevier B.V.)

Interferon (IFN)-α is able to stimulate many cellular genes and inhibit the replication of various viruses. However, it is unknown whether some IFN-stimulated genes (ISGs) specifically inhibit hepatitis B virus (HBV) replication. Therefore, we attempted to identify ISGs with antiviral activities against HBV. Knockdown of IFN-induced proteins with tetratricopeptide repeats 1 and 2 (IFIT1 and IFIT2) in HepG2.2.15 led to markedly increased HBV replication. Consistently, this effect was verified by transient transfection with a replication-competent HBV clone in HepG2 and Huh7. However, IFN-α stimulation could override the knockdown by siRNAs and enhance the expression of IFIT1 and IFIT2, leading to reduced HBV replication. Silencing of IFIT1 or IFIT2 decreased the expression of the corresponding genes while other ISGs like MxA were not affected. Northern blot analysis showed that IFIT1 and IFIT2 knockdown slightly increased the levels of HBV 3.5, 2.4 and 2.1 kb transcripts, while IFIT1 and IFIT2 overexpression did not change their levels. Consistently, the reporter assays with HBV promoters demonstrated that IFIT1 and IFIT2 differentially but only modestly regulated HBV promoter activity. Thus, IFIT1 and IFIT2 contribute significantly to the regulation of HBV replication, likely at both transcriptional and posttranscriptional steps., (Copyright © 2013 S. Karger AG, Basel)

Published studies have described a strong association with a single-nucleotide polymorphism (SNP) in the inosine triphosphate pyrophosphatase (ITPA) gene and ribavirin (RBV)-induced hemolytic anemia in HCV-infected patients receiving pegylated interferon (pegIFN) and RBV. This study sought to evaluate the effect of these polymorphisms on anemia, hemoglobin reduction, HCV kinetics, and treatment outcomes. Sixty-three patients coinfected with HIV and HCV and 58 patients infected with HCV only were treated with pegIFN/RBV were genotyped using the ABI TaqMan allelic discrimination kit for the 2 ITPA SNP variants rs1127354 and rs7270101. A composite variable of ITPA deficiency using both SNPs was created as previously reported. Statistical analysis was performed using Mann-Whitney test or Chi square/Fishers exact test for categorical data and mixed model analysis for multiple variables. Thirty-five patients (30%) were predicted to have reduced ITPA activity. ITPA deficiency was found to be protective against the development of hemoglobin reduction >3 g/dl over the course of treatment. The rates of hemoglobin reduction >3 g/dl decreased in correlation with the severity of ITPA deficiency. ITPA deficiency was associated with slower hemoglobin decline early in treatment (week 4, P = 0.020) and rapid virologic response (RVR) at week 4 (P = 0.017) in patients coinfected with HIV and HCV. ITPA polymorphisms are associated with hemoglobin decline and in patients coinfected with HIV and HCV it is also associated with early virologic outcomes. Determination of ITPA polymorphisms may allow prediction of RBV-induced anemia and earlier initiation of supportive care to ensure optimal therapeutic outcomes., (Published 2012. This is a US Government work and as such is in the public domain in the United States of America.)

Interferons (IFNs) are widely used in therapy for viral, neoplastic, and inflammatory disorders, but clinical response varies among patients. The biological basis for variable clinical response is not known. We determined the primary molecular response to IFN-beta (IFN-beta) injections in 35 treatment-naïve multiple sclerosis (MS) patients using a customized cDNA macroarray with 186 interferon-stimulated genes (ISGs). Our results revealed striking interindividual heterogeneity, both in the magnitude as well as the nature of the primary molecular response to IFN-beta injections. Despite marked between-subject variability in the molecular response, responses within individual subjects were stable over a 6-month interval. Our data suggest that clinical response to IFN-beta therapy for MS differs among patients because of qualitative rather than quantitative variability in the primary molecular response to the drug.

We report the case of a 74-year-old female with an extreme picture of melanosis coli of the whole colon after chronic use of anthraquinone laxatives for the treatment of constipation over many decades. Endoscopic work-up revealed an impressive deep black pigmentation of the whole colon mucosa which could be verified by histopathology as a widespread lipofuscin granulation. In addition, various adenomas but no colorectal carcinoma could be detected. The term melanosis coli describes a brown or black pigmentation of the colonic mucosa. Induction of melanosis coli by anthraquinone laxatives and their derivatives can be regarded as verified. The question if melanosis coli predisposes for colorectal neoplasia is discussed controversially. Based on the current literature, an association of melanosis coli between colorectal adenomas, but not colorectal carcinomas, is under discussion but the mechanisms to effect the development of colorectal neoplasia are not completely understood. Considering our case and the current scientific backround, we conclude that due to pharmaceutical side effects of anthraquinone derivatives such as electrolytic shift and water loss in addition to the risk of developing melanosis coli, anthraquinone laxatives should not be used for long-term therapy of constipation.

Small interfering RNA (siRNA) has been shown to be active to inhibit the hepatitis B virus gene expression and replication in transient and stable transfection systems. Here in primary hepatocytes prepared from naturally woodchuck hepatitis virus (WHV)-infected woodchucks, four siRNAs targeting the WHV preS1, S, C, and X region led to a depletion of WHV transcripts and replicative intermediates with different kinetics and a decreased production of viral particles. Two siRNAs targeting WHV S and X region had the highest efficacy to deplete 70% of WHV transcripts and replicative intermediates. In addition, siRNA-mediated suppression of WHV enhanced the expression of cellular genes like MxA and MHC I. Specific siRNAs are able to inhibit the hepadnaviral replication and enhance the expression of cellular genes relevant for antiviral actions. Thus, siRNAs might be useful as novel antiviral agents for the treatment of chronic HBV infection.

Background: In chronic hepatitis B patients undergoing therapy with LAM or ADV, viral breakthrough is possible due to the emergence of drug resistance. LAM resistant HBV strains are susceptible to ADV, while ADV resistant mutants remain sensitive to LAM., Case Report: A male patient with HBV-related cirrhosis developed viral breakthrough (HBV DNA>1.8 x 106 IU/ml) after 4 1/2 years of treatment with LAM, and therapy was switched to ADV (10 mg/d). After three months, HBV remained highly replicative without any changes of ALT values, and ADV dose was increased (20 mg/d). Because of unchanged VL sequence analysis was performed three months later, which showed the mutation (rtS219A) and the concomitant mutation (sS210R) and 2 mutations in core promoter region (A1762T), (G1764A). During the sixth month of ADV monotherapy the patient developed liver failure. After administration of TDF plus LAM, HBV DNA became undetectable within 39 days. At day 41, the patient underwent OLT. TDF plus LAM were well tolerated, and the patient maintained undetectable HBV DNA levels, and in addition to HBIG a sustained HBsAg negative status over twenty-eight months post OLT., Conclusion: TDF plus LAM is a safe drug combination in case of viral breakthrough during LAM treatment and subsequent primary non-response to ADV. High VL persisting for >or= 6 months of continuous antiviral treatment may indicate drug resistance. Especially in cirrhotic patients with LAM resistance, "add on" of a nucleotide analogue is the right therapeutic strategy even before viral breakthrough gets apparent.

Psychosomatic assessment for living liver donors for adults has as yet not been described in detail. Between August 1998 and September 2003, 205 donor candidates were admitted for psychosomatic evaluation; 13.2% of whom (N=27) were excluded. Mental disturbances, especially when accompanied with social stress, were frequent reasons for exclusion. Selected donors had higher social status with regard to education and employment rate than other donor candidates. Candidates with better psychosocial resources are selected in the psychosomatic evaluation, and this contributes to the good psychosocial outcome of living donors.

Background & Aims: Endoscopic treatment of biliary strictures after liver transplantation is a therapeutic challenge. In particular, outcomes of endoscopic therapy of biliary complications in the case of duct-to-duct anastomosis after living related liver transplantation are limited. The aim of this study was to evaluate the feasibility and success of an endoscopic treatment approach to posttransplant biliary strictures (PTBS) after right-sided living donor liver transplantation (RLDLT) with duct-to-duct anastomosis., Methods: Ninety patients who received adult-to-adult RLDLT in our center were screened retrospectively with respect to endoscopic treatment of PTBS. Therapy was judged as successful when cholestasis parameters returned to normal and bile duct narrowing was reduced significantly after the completion of therapy., Results: Forty of 90 RLDLT patients received duct-to-duct anastomosis, 12 (30%) showed PTBS. Seven of 12 patients were treated successfully by endoscopy; the remaining 5 patients were treated primarily by surgery. Most patients were treated by balloon dilatation followed by insertion of endoprostheses. A median of 2.5 dilatation sessions were necessary and the median treatment duration was 8 months. One patient developed endoscopy-treatable recurrent stenosis, no surgical intervention was necessary. Mild pancreatitis occurred in 7.9% and cholangitis in 5.3% of the procedures. One minor bleeding episode occurred during sphincterotomy. Bleeding was managed endoscopically., Conclusions: Endoscopic therapy of adult-to-adult right living related liver transplantation with duct-to-duct anastomosis is feasible and frequently is successful. The duct-to-duct anastomosis offers the possibility of endoscopic treatment. Endoscopic treatment of posttransplant biliary strictures is safe, with a low specific complication rate.

Objective: To investigate the expression profile of genes which are involved in IFN antiviral activity and IFN signal transduction pathway in Hep G2 and HepG2.2.15 cells., Methods: Genes of interest were selected from the UniGene database (http://www.ncbi.nlm.gov/UniGene/Hs.Home.html). The 5'IMAGE clones with 0.5-0.8 kb length were chosen and ordered from RZPD company. The cDNA inserts were amplified by PCR and then were spotted onto the Hybond-N+ membranes. The membranes were denatured and neutralized for Macroarray analysis. HepG2.2.15 and Hep G2 cells were treated without or with IFN-alpha for 6 h, and the total cellular RNA was isolated using Trizol Reagent. Radio-labelled cDNA was generated from 20 microgram of RNA by reverse transcription using 360 units of reverse transcriptase in the presence of 30 microCi of alpha-32P dCTP. Hybridization was performed between 32P-labelled cDNA and membrane arrays. The membranes were then scanned, and the intensity of autoradiographic spots was quantitated by Cyclone Storage Phosphor System. The images were subsequently analysed by the OptiQuant Imager Analysis Software and converted into digital data., Results: The authors found that just partially IFN-inducible genes were expressed in Hep G2 and HepG2.2.15 cells, and the majority of IFN-inducible genes was lowly responsive or non-responsive to IFN-a treatment. Some interferon-stimulated genes (ISGs) were inhibited or blocked, especially in HepG2.2.15 cells. Interestingly, the authors found that the IFN signal transduction pathway (Jak-STAT) was intact and unimpaired in HepG2.2.15 cells., Conclusion: Differential gene expression profiles in response to IFN were found between Hep G2 and HepG2.2.15 cells.

Aim: To evaluate the diagnostic value of different indirect methods like biochemical parameters, ultrasound (US) analysis, CT-scan and MRI/MRCP in comparison with endoscopic retrograde cholangiography (ERC), for diagnosis of biliary complications after liver transplantation., Methods: In 75 patients after liver transplantation, who received ERC due to suspected biliary complications, the result of the cholangiography was compared to the results of indirect imaging methods performed prior to ERC. The cholangiography showed no biliary stenosis (NoST) in 25 patients, AST in 27 and ITBL in 23 patients., Results: Biliary congestion as a result of AST was detected with a sensitivity of 68.4% in US analysis (specificity 91%), of 71% in MRI (specificity 25%) and of 40% in CT (specificity 57.1%). In ITBL, biliary congestion was detected with a sensitivity of 58.8% in the US, 88.9% in MRI and of 83.3% in CT. However, as anastomotic or ischemic stenoses were the underlying cause of biliary congestion, the sensitivity of detection was very low. In MRI detected the dominant stenosis at a correct localization in 22% and CT in 10%, while US failed completely. The biochemical parameters, showed no significant difference in bilirubin (median 5.7; 4,1; 2.5 mg/dL), alkaline phosphatase (median 360; 339; 527 U/L) or gamma glutamyl transferase (median 277; 220; 239 U/L) levels between NoST, AST and ITBL., Conclusion: Our data confirm that indirect imaging methods to date cannot replace direct cholangiography for diagnosis of post transplant biliary stenoses. However MRI may have the potential to complement or precede imaging by cholangiography. Optimized MRCP-processing might further improve the diagnostic impact of this method.

Alpha/beta interferon (IFN-alpha/beta) is a key mediator of innate antiviral responses but has little effect on the established replication of dengue viruses, which are mosquito-borne flaviviruses of immense global health importance. Understanding how the IFN system is inhibited in dengue virus-infected cells would provide critical insights into disease pathogenesis. In a recent study analyzing the ability of individual dengue virus-encoded proteins to antagonize the IFN response, nonstructural (NS) protein 4B and possibly NS2A and NS4A were identified as candidate IFN antagonists. In monkey cells, NS4B appeared to inhibit both the IFN-alpha/beta and IFN-gamma signal transduction pathways, which are distinct but overlapping (J. L. Munoz-Jordan, G. G. Sanchez-Burgos, M. Laurent-Rolle, and A. Garcia-Sastre, Proc. Natl. Acad. Sci. USA 100:14333-14338, 2003). For this study, we examined the effects of dengue virus on the human IFN system, using cell lines that were stably transfected with self-replicating subgenomic dengue virus RNA (replicons) and that expressed all of the dengue virus nonstructural proteins together. We show here that in replicon-containing cells dengue virus RNA replication and the replication of encephalomyocarditis virus, an IFN-sensitive virus, are resistant to the antiviral effects of IFN-alpha. The presence of dengue virus replicons reduces global IFN-alpha-stimulated gene expression and specifically inhibits IFN-alpha but not IFN-gamma signal transduction. In cells containing replicons or infected with dengue virus, we found reduced levels of signal transducer and activator of transcription 2 (STAT2), which is a key component of IFN-alpha but not IFN-gamma signaling. Collectively, these data show that dengue virus is capable of subverting the human IFN response by down-regulating STAT2 expression.

Background: It has been shown that interaction of anti-PR-3 antibodies with human endothelial cells (EC) leads to an activation of EC in vitro, i.e. induction of adhesion molecules like E-selectin, VCAM-1 and tissue factor. The aim of this study was to investigate the effect of anti-PR-3 antibodies on endothelial IL-8 expression., Materials and Methods: EC were cultured in 96-well plates and stimulated with TNF-alpha and IL-1beta for 1 h to induce membrane expression of endothelial PR-3. Anti-PR-3 antibodies were purified from sera from patients with clinically active Wegener's granulomatosis. Purified anti-Ro, anti-centromere, anti-dsDNA antibodies and a monoclonal anti-PR-3 antibody (WGM2) served as controls. Induction of IL-8 mRNA was detected by RT-PCR. IL-8 was measured in the supernatant of EC by ELISA. In addition, induction of NFkappaB was investigated by PAGE of nuclear extracts of EC and Western blot with ab against p65., Results: In contrast to controls, interaction of anti-PR-3 antibodies (patients and WGM2) with cytokine activated EC led to the highest amount of IL-8 synthesis. Priming of EC with cytokines alone induced a markedly lower IL-8 level. The lowest levels of IL-8 could be measured after incubation of unprimed EC with anti-PR-3 antibodies. Anti-PR-3 antibody induced endothelial IL-8 expression could be inhibited by cycloheximide. In addition, we established that the activation of NF-kappaB is critically involved in anti-PR-3 antibody induced endothelial IL-8 production., Conclusion: In summary, we were able to show that anti-PR-3 antibodies induce endothelial IL-8 synthesis by activating NF-kappaB. As IL-8 represents a powerful neutrophil chemoattractant, our data provide further evidence for a direct pathogenic effect of anti-PR-3 antibodies in ANCA related diseases.

Methotrexate (MTX) is an effective immunosuppressive agent in various chronic inflammatory diseases such as rheumatoid arthritis (RA). However, its mechanisms of action are only partially understood. In this study, we assessed the effects of MTX on the differentiation of peripheral blood (PB) CD4+CD45RA 'naive' and CD4+CD45RO 'memory' T cells from healthy controls and patients with RA. Accordingly, purified T cells were primed and restimulated in vitro via the T cell receptor (TCR) in the presence of IL-2 to generate effector T cells secreting large amounts of Th1 and Th2 cytokines. We observed that low doses of MTX strongly suppress TNF and to a lesser extent interferon-gamma (IFN-gamma) production by T cells from both healthy donors and RA patients when present during T cell priming via the TCR. Similar data were obtained for TCR-primed synovial fluid mononuclear cells in RA. In contrast, production of IL-4 by TCR-primed CD45RA T cells was significantly increased upon MTX treatment. Interestingly, MTX did not enhance IL-4 production when present during restimulation of effector CD45RO T cells, although it still suppressed TNF production. The results indicate that MTX effects depend on the stage of T cell activation and identify TNF production by TCR-primed T lymphocytes as a target for low-dose MTX treatment in RA. These findings could explain the delayed clinical effects of MTX and may contribute to its potent anti-inflammatory and immunoregulatory properties.

The initial stage of Wegener's Granulomatosis (WG) is often marked by symptoms of infection and it has been postulated that a bacterial infection could be the aetiologic factor of this disease. The objective of our work was to investigate T-cell-mediated immunity in WG by testing proliferative responses on bacterial antigens and particularly Staphylococcus aureus. We investigated the bulk proliferative response of peripheral blood lymphocytes (PBL) from patients with clinically active WG to gram-positive bacteria and purified proteinase 3 (PR-3), the major target antigen of c-ANCA. We generated S. aureus-specific PBL-derived T-cell lines and T-cell clones (TLC). In two WG patients 27 TLC were characterized in terms of reactivity to bacterial antigens/PR-3, phenotype, HLA class II restriction and pattern of cytokine secretion. Compared to coagulase-negative Staphylococci and beta-haemolytic Streptococci A, reactivity to S. aureus was significantly increased in all patients with WG. Using purified PR-3, we found a PBL proliferation in five out of 25 WG patients. The TLC were S. aureus-specific and did not cross-recognize Streptococci or coagulase-negative Staphylococci. The S. aureus-specific TLC were of the alphabeta-TCR+ CD4+ phenotype and HLA-DR-restricted. These TLC predominantly showed a Th2-type of cytokine secretion. Interestingly, seven of the S. aureus-reactive TLC also recognized the PR-3 antigen. From these data we conclude that Staphylococci-specific HLA-DR-restricted CD4+ T cells may play a key role in the initial triggering of immune responses in WG.

Because cellular and humoral immune responses against the hepatitis B virus (HBV) surface antigen (HBs) might be crucial to overcome HBV infection, HBs-specific B- and T-cell responses of HBV patients and HBs vaccine recipients were analyzed quantitatively and functionally. In patients with acute hepatitis B (AHB), transient high anti-HBs-secreting B-cell frequencies were observed early after clinical onset, whereas 1 patient who probably developed chronic infection and chronic HBV carriers had absent or weak B- and T-cell responses. In HBs vaccine recipients, maximal HBs-specific B- and T-cell responses were detected after the first injection that decreased gradually before anti-HBs antibodies appeared in serum. Years after vaccination, anti-HBs-secreting B cells were enriched in the bone marrow. After in vitro stimulation with HBsAg, peripheral blood mononuclear cells (PBMC) of only 1 of 5 acute and 1 of 6 chronic HBV patients, but of all 6 vaccine recipients, secreted varying amounts of interferon gamma (IFN-gamma), but no interleukin-4 (IL-4) or IL-5. Furthermore, the addition of IFN-gamma, but not of IL-2, -4, -12, or IFN-alpha, resulted in strong increases of anti-HBs-secreting B cells in vaccine recipients and chronic carriers. In conclusion, circulating anti-HBs-secreting B cells were significantly higher in early acute hepatitis B or early after HBs vaccination than in chronic hepatitis B and decreased in the follow-up as a result of compartmentalization to lymphoid tissues. Release of IFN-gamma by antigen-stimulated T cells might be critical for anti-HBs formation.

Aims/background: CD4+ T-helper cell (Th) responses to hepatitis B virus (HBV) core antigen (HBc) are increased during exacerbations in acute and chronic hepatitis B (AHB, CHB) and might influence the induction of CD8+ cytotoxic T lymphocytes (CTL) that are important for viral clearance., Methods: HBc-specific proliferative responses and cytokine release of blood mononuclear cells (PBMC) were studied in patients with AHB or CHB, as well as responders and non-responders to interferon-alpha treatment (IFN-R, IFN-NR), by [3H]-thymidine-uptake, enzyme-linked immunosorbent assay (ELISA) and Elispot assay and were compared to peptide HBc18 27-specific CTL precursor frequencies among CD8+ T cells derived from HLA-A2+ patients., Results: HBc-specific proliferative PBMC responses and Th frequencies were significantly increased in AHB patients compared with untreated CHB patients. PBMC derived from IFN-R showed stronger cellular responses than IFN-NR. Stimulated PBMC from all patient groups secreted significantly more IFN-gamma than IL-4 indicating Th1/Th0 cell responses. Furthermore, in AHB and IFN-R patients, high peptide HBc18-27-specific CTL precursor frequencies closely correlated with strong HBc-specific Th responses, whereas in untreated CHB and IFN-NR patients lower CTL frequencies were observed without correlation to Th activities., Conclusions: HBV core-specific Th-cell responses appeared to support efficient CTL induction in patients with viral clearance, whereas in chronic HBV carriers quantitatively insufficient Th and CTL responses were observed. This observation could be important for future therapeutic strategies.

The cytokine profile of tumour reactive T cells is likely to play a central role in their function. However, little is known about how cytokine patterns of tumour reactive T cells can be regulated. Here, the authors investigated the influence of exogenous regulatory cytokines in addition to interleukin-2 (IL-2) on cytokine patterns and the proliferation of T cells recognizing an autologous sarcoma cell line. In this system, IL-4 and IL-12 showed the most polarizing influences on tumour reactive T cells. Exogenous IL-4 induced a predominant production of IL-4 while decreasing the interferon-gamma (IFN-gamma) and IL-10 production by tumour reactive T cells. It also stimulated the growth of tumour reactive CD4+ T cell clones. In contrast, IL-12 substantially increased the production of IL-10 and IFN-gamma. This was accompanied by a growth inhibition of tumour reactive T cells. The growth of CD4+ tumour reactive T cells was also suppressed by exogenous IL-10. This study shows that cytokine patterns and proliferation tumour reactive T cells can be significantly influenced by exogenous cytokines and confirms the hypothesis of a negative feedback loop of IL-12 by the induction of IL-10 in the context of human tumour reactive T cells.

Sarcoidosis is a systemic, granulomatous disorder with a high rate of spontaneous remission indicating the presence of antiinflammatory mechanisms. Antiinflammatory mediators such as interleukin-10 (IL-10) and transforming growth factor-beta (TGF-beta) should be able to induce spontaneous remission of sarcoidosis. By measuring the release of both mediators in culture supernatants of bronchoalveolar lavage (BAL) cells, we investigated their relevance in the spontaneous remission of sarcoidosis. No spontaneous IL-10 release was observed by BAL cells of sarcoid patients. In supernatants of BAL cells of seven patients found retrospectively to be free of any interstitial lung disease, we found 612 +/- 261.2 pg/ml (mean +/- SEM) TGF-beta. TGF-beta release was recorded in 20 of 39 patients with active disease. Patients with active disease without TGF-beta release in BAL cell culture either required therapy (n = 21; 677 +/- 159 pg/ml) or showed evidence of persisting disease (n = 6; 762 +/- 419 pg/ml). Patients with active disease without indications for therapy and with significantly increased TGF-beta release (n = 12; 1,422 +/- 215 pg/ml; p < 0.004 in all comparisons) had a spontaneous remission within 6 mo. Increased TGF-beta release (1,560 +/- 353 pg/ml) was observed in five of five patients receiving therapy. We conclude that TGF-beta is a regulator of the inflammatory process in sarcoidosis.

CD4+ T cells play an important role for tumor immunity in animal tumor models, yet there are few reports about the role of CD4+ HLA class II-restricted T cells in the immune response against human tumors. Against a human sarcoma exclusively CD4+, T cell clones could be established. These T cell clones were cytotoxic and secreted TNF and additional cytokines in response to the IFN-gamma-treated, HLA class II-positive autologous sarcoma cells. Several Ags were recognized by representative T cell clones: an Ag presented by HLA-DR and specific for the sarcoma; Ags presented by both HLA-DR alleles of the sarcoma, HLA-DR4 and -15, and shared by allogenic HLA-DR matched cell lines of different tissue lineages, including B cell blasts; and a sarcoma Ag presented by HLA-DP or DQ. Cytokine profiles of sarcoma-reactive T cell clones were dependent on the cytokine environment present during establishment of the T cell clones. The addition of exogenous IL-4 shifted the cytokine patterns of sarcoma-reactive T cell clones from Th1-like patterns to Th0/Th2-like patterns and decreased IL-10 production. TNF, IFN-gamma, IL-4, and supernatants of T cell clones induced HLA-DR expression on the sarcoma cells and, thus, were able to enhance Ag presentation. This autologous T cell response to a human sarcoma represents a new model for HLA class II-restricted T cell responses to human tumors.

We evaluate the influence of IL-4, IL-10 and TGF-beta upon the release of IL-1 alpha, tumor necrosis factor-alpha (TNF-alpha), and IL-6 by lipopolisaccharide (LPS, 1 microgram/ml) stimulated alveolar macrophages (AM). IL-4 reduced TNF-alpha release, in a dose dependent manner, to 62% and IL-1 alpha release to 42% of LPS-stimulated AM without IL-4. IL-6 release was also suppressed (61%), however, with a biphasic dose response curve. IL-10 suppressed LPS induced release of IL-alpha and TNF-alpha to approximately 50% of control without affecting IL-6 release. When used at high concentrations, TGF-beta achieved moderate reductions of TNF-alpha and IL-1 alpha release. Low concentrations of LPS (0.1 microgram/ml), allowed a dose-dependent TGF-beta-induced suppression of TNF-alpha-release to approximately 80% of control. The combinations of IL-4 and IL-10 was more effective in suppressing IL-6 release, although the suppression was only weak. Other combinations of cytokines revealed no synergistic inhibitory activities. Our data demonstrate that IL-1 alpha and TNF-alpha release by AMs is down regulated by IL-4, IL-10, and TGF-beta. IL-6 release, however, can only be suppressed to some extent by a combination of IL-4 and IL-10.

Endothelin-1 is known to possess various biological properties. In the present study we have investigated the effects of Endothelin-1 (Et-1) on the expression of adhesion molecules ICAM-1, VCAM-1 and CD-44 by fibroblast-like synoviocytes. Cultured fibroblast-like synoviocytes were treated with Et-1 in the absence or presence of C1306, a specific endothelin-A-receptor antagonist. Cell surface expression of ICAM-1, VCAM-1 and CD-44 was determined by immunofluorescence studies, Cyto-ELISA and FACS-analysis. ICAM-1, VCAM-1 and CD-44 were constitutively expressed on cultured FLS. After incubation with Et-1 the expression of ICAM-1, VCAM-1 and CD-44 increased. The level of expression of adhesion molecules after Et-1 stimulation was similar to cytokine mediated effects (IL-1 beta, TNF-alpha). IL-1 beta showed the strongest stimulatory effect on the expression of ICAM-1, TNF-alpha preferentially induced the expression of VCAM-1 and CD-44, and Et-1 strongly stimulated the upregulation of CD-44 expression. In addition, Et-1 enhanced significantly the IL-1 beta mediated upregulation of VCAM-1 expression, whereas TNF-alpha mediated expression of VCAM-1 was downregulated by Et-1. Furthermore, the Et-1 induced expression of adhesion molecules on FLS was mediated via the endothelin-A-receptor (EtA-receptor), since C-1306, a selective endothelin-A-receptor antagonist, could block this effect. These results indicate that Et-1 has stimulating effects on FLS in vitro. The expression of adhesion molecules can be upregulated by Et-1 similar to proinflammatory cytokines. The modulating effect of Et-1 can be inhibited by the pretreatment with a selective EtA-receptor antagonist. Thus, Et-1 may have immunoregulatory functions in the recruitment of cells infiltrating the inflamed tissue.