[Autoimmune hepatitis-standard and second-line therapy]. / Autoimmunhepatitis ­ Standard- und Zweitlinientherapie.

BACKGROUND: Autoimmune hepatitis (AIH) is a rare chronic inflammatory liver disease. Like other autoimmune diseases, the incidence continues to rise and, if left untreated, it has a detrimental natural course. AIM: Description of the current etiology, diagnosis and management of AIH. MATERIALS AND METHODS: Summary of current national as well as international guidelines and highlighting recently published studies. RESULTS AND CONCLUSION: In addition to autoantibody testing and liver histology, the diagnosis of AIH is based on the exclusion of other relevant liver diseases. The standard of care with prednisolone ± azathioprine, which has been well established for decades, significantly improves long-term survival. The therapeutic aim is the complete normalization of aminotransferases and immunoglobulin G. Budesonide, an alternative topical steroid with fewer systemic side effects, was approved for AIH therapy in non-cirrhotic patients in 2011. The therapeutic goal of complete biochemical remission is achieved in about 80% of patients and liver transplantation is rarely necessary. The majority of patients require life-long immunosuppressive therapy because of high relapse rates after discontinuation of immunosuppressants. Currently used second-line therapies are based on uncontrolled monocentric studies with single substances. Therefore, clear recommendations from international expert groups are lacking. These second-line therapies are not approved by the regulatory authorities such as the Food and Drug Administration or the European Medicines Agency.

Hepatic Infiltrates in Operational Tolerant Patients After Liver Transplantation Show Enrichment of Regulatory T Cells Before Proinflammatory Genes Are Downregulated.

Immunosuppression can be discontinued from selected and stable patients after liver transplantation resulting in spontaneous operational tolerance (SOT), although the underlying mechanisms remain elusive. Thus, we analyzed serial liver biopsy specimens from adult liver recipients enrolled in a prospective multicenter immunosuppression withdrawal trial that used immunophenotyping and transcriptional profiling. Liver specimens were collected before the initiation of weaning, at the time of rejection, or at 1 and 3 years after complete drug discontinuation. Unexpectedly, the tolerated grafts developed portal tract expansion with increased T cell infiltration after immunosuppression withdrawal. This was associated with transient and preferential accumulation of CD4(+) FOXP3(+) cells and a trend toward upregulation of immune activation and regulatory genes, without signs of rejection. At the same time, no markers of endothelial damage or activation were noted. Portal infiltrates persisted at 3 years but were characterized by decreased expression of genes associated with chronic immunological damage. Further, SOT was not associated with a progressive liver fibrosis up to 5 years. These data suggest that SOT involves several mechanisms: a long-lasting local immune cell persistence with a transient regulatory T cells accumulation followed by a downregulation of immune-activated genes over years. These results have important implications for designs and follow-up of weaning trials.

[Chronic rejection: Differences and similarities in various solid organ transplants]. / Chronische Abstoßung: Unterschiede und Ähnlichkeiten bei verschiedenen soliden Organtransplantationen.

In this paper, chronic rejections after transplantation of the lungs, heart, liver, and kidney are described. Chronic allograft dysfunction (CAD) plays an important role in all of these transplantations and has a significant influence on patient survival. The pathophysiological reasons for CAD varies greatly in the various organs.Chronic lung allograft dysfunction (CLAD) is the most important determinant of survival and quality of life after lung transplantation. Diagnosis is based on lung function, especially forced expiratory flow in 1 s (FEV1) decline. Prevention, early detection, and rapid treatment are extremely important. Azithromycin and extracorporeal photopheresis are commonly used for treatment because they usually positively influence the progression of lung remodeling.The expression for chronic rejection of the heart is cardiac allograft vasculopathy (CAV). Immunological and nonimmunological factors are important for its development. Due to limited therapeutic options, prevention is of utmost importance (administration of mTOR inhibitors and minimizing cardiovascular risk factors).The mid- and long-term survival rates after liver transplantation have hardly changed in recent decades, which is an indication of the difficulty in diagnosing chronic graft dysfunction. Chronic ductopenic rejection accounts for a small proportion of late graft dysfunction. Idiopathic posttransplant hepatitis and de novo autoimmune hepatitis are important in addition to recurrence of the underlying disease that led to transplantation.Chronic allograft nephropathy is the result of severe rejection which cumulates in increasing fibrosis with remodeling. The earliest possible diagnosis and therapy is currently the only option. Diagnosis is based on evidence of donor-specific antibodies and histological findings.

Naive tumour-specific CD4+ T cells were efficiently primed in acute lymphoblastic leukaemia.

The recognition and neutralization of tumour cells is one of the big challenges in immunity. The immune system has to recognize syngeneic tumour cells and has to be primed and respond in an adequate manner. Priming of a leukaemia-specific immune response is a crucial step in tumour immunology that can mislead to tumour tolerance either by T cell ignorance, deletion or Treg induction. To resemble the situation of acute lymphoblastic leukaemia (ALL) in patients, we used the murine BALB/c model with syngeneic BM185 tumour cells. We established a tumour cell line that expresses the neo-antigen ovalbumin (BM185-OVA/GFP) to allow the application of T cell receptor transgenic, antigen-specific CD4(+) T cells. Here, we demonstrate that effective anti-ALL immunity can be established by in vivo priming of CD4(+) T cells that is sufficient to differentiate into effector cells. Yet they failed to control tumour alone, but initiated a Th1 response. An efficient tumour clearance was dependent on both antigen-specific CD4(+) T cells and CD8(+) effector T cells from the endogenous repertoire. The tolerogeneic milieu was characterized by increased Tregs numbers and elevated IL-10 level. Tregs hamper effective antitumour immune response, but their depletion did not result in reduced tumour growth. In contrast, neutralization of IL-10 improved median mouse survival. Future therapies should focus on establishing a strong CD4+ T cells response, either by adjuvant or by adoptive transfer.

Enrichment of regulatory T cells in acutely rejected human liver allografts.

Acute cellular rejection (ACR) occurs frequently after liver transplantation and can usually be controlled. Triggering of allospecific immune responses and lack of immunoregulation are currently suggested as a cause of ACR, but there are no investigations of intrahepatic immune responses during ACR. Therefore we prospectively analyzed the intrahepatic T cell infiltration pattern in correlation to the severity of ACR in a cohort of patients with graft hepatitis (n = 151). While CD4(+) cells dominated the portal infiltrates in mild-moderate ACR, CD8(+) cells prevailed in severe ACR. Furthermore portal CD8(+) and not CD4(+) infiltration correlated with serum transaminases and with the likelihood of subsequent ACRs. Surprisingly, the rise of portal effector T cells density during ACR was surpassed by the increase in portal infiltration of regulatory T cells by a factor of two. Thus ACRs rather showed an increase and not a lack of regulation, as was suggested by analysis of peripheral blood mononuclear cells. Despite the pattern of enhanced immunoregulation, patients with severe ACR had a higher risk for subsequent rejections and showed a trend to a reduced survival. Thus, patients with severe rejections might need a modification of their immunosuppression to improve prognosis.

Chronic hepatitis e in heart transplant recipients.

Chronic courses of hepatitis E virus (HEV) infections have been described in immunosuppressed patients. We aimed to study the role of HEV infections in heart transplant recipients (HTR). 274 HTR were prospectively screened for HEV infection using an anti-HEV-IgG ELISA and HEV-PCR. In addition, 137 patients undergoing cardiac surgery (non-HTR) and 537 healthy subjects were studied cross-sectionally. The anti-HEV-IgG seroprevalence was 11% in HTR, 7% in non-HTR and 2% in healthy controls (HTR vs. healthy controls p<0.0001; non-HTR vs. healthy controls p<0.01). Anti-HEV tested positive in 4.0% in control cohorts of other immunocompromised patients (n = 474). Four HTR (1.5%) were chronically infected with HEV as shown by HEV-PCR and all four patients had liver transaminases of >200 IU/L and histological or clinical evidence of advanced liver disease. In three patients ribavirin treatment was successful with a sustained biochemical and virological response while treatment failed in one cirrhotic patient after ribavirin dose reduction. Heart transplant recipients and patients undergoing cardiac surgery have an increased risk for HEV infections. Chronic hepatitis E may explain elevated liver enzymes in heart transplant recipients. Treatment of HEV infection with ribavirin is effective but the optimal dose and duration of ribavirin therapy remains to be determined.

State-of-the art comparability of corrected emission spectra. 1. Spectral correction with physical transfer standards and spectral fluorescence standards by expert laboratories.

The development of fluorescence applications in the life and material sciences has proceeded largely without sufficient concern for the measurement uncertainties related to the characterization of fluorescence instruments. In this first part of a two-part series on the state-of-the-art comparability of corrected emission spectra, four National Metrology Institutes active in high-precision steady-state fluorometry performed a first comparison of fluorescence measurement capabilities by evaluating physical transfer standard (PTS)-based and reference material (RM)-based calibration methods. To identify achievable comparability and sources of error in instrument calibration, the emission spectra of three test dyes in the wavelength region from 300 to 770 nm were corrected and compared using both calibration methods. The results, obtained for typical spectrofluorometric (0°/90° transmitting) and colorimetric (45°/0° front-face) measurement geometries, demonstrated a comparability of corrected emission spectra within a relative standard uncertainty of 4.2% for PTS- and 2.4% for RM-based spectral correction when measurements and calibrations were performed under identical conditions. Moreover, the emission spectra of RMs F001 to F005, certified by BAM, Federal Institute for Materials Research and Testing, were confirmed. These RMs were subsequently used for the assessment of the comparability of RM-based corrected emission spectra of field laboratories using common commercial spectrofluorometers and routine measurement conditions in part 2 of this series (subsequent paper in this issue).

State-of-the art comparability of corrected emission spectra. 2. Field laboratory assessment of calibration performance using spectral fluorescence standards.

In the second part of this two-part series on the state-of-the-art comparability of corrected emission spectra, we have extended this assessment to the broader community of fluorescence spectroscopists by involving 12 field laboratories that were randomly selected on the basis of their fluorescence measuring equipment. These laboratories performed a reference material (RM)-based fluorometer calibration with commercially available spectral fluorescence standards following a standard operating procedure that involved routine measurement conditions and the data evaluation software LINKCORR developed and provided by the Federal Institute for Materials Research and Testing (BAM). This instrument-specific emission correction curve was subsequently used for the determination of the corrected emission spectra of three test dyes, X, QS, and Y, revealing an average accuracy of 6.8% for the corrected emission spectra. This compares well with the relative standard uncertainties of 4.2% for physical standard-based spectral corrections demonstrated in the first part of this study (previous paper in this issue) involving an international group of four expert laboratories. The excellent comparability of the measurements of the field laboratories also demonstrates the effectiveness of RM-based correction procedures.

Donor-specific regulatory T cells generated on donor B cells are superior to CD4+CD25high cells in controlling alloimmune responses in humanized mice.

CD4(+)CD25highFOXP3(+) regulatory T cells (Tregs) can control allospecific immune responses in vitro and in titrated lymphopenic transplantation models. However, under non-lymphopenic conditions, as seen in patients with autoimmune diseases or after organ transplantation, polyspecific Tregs so far have been largely ineffective to control immune responses in animal models. Yet currently polyspecific CD4(+)CD25high Tregs are being tested in clinical trials. Donor materials are usually limited for the generation of donor-specific Tregs. Herein we have developed a method to produce large quantities of activated donor B cells by stimulation of donor peripheral blood mononuclear cells with 3T3 fibroblasts expressing CD40L. These activated donor B cells are potent stimulators of CD4(+)CD25high Tregs, which were expanded efficiently to inhibit an allo-MLR in donor-specific fashion. They were far more potent in inhibiting alloimmune responses in humanized mice compared with the polyspecific CD4(+)CD25high Tregs. Generation of donor-specific Tregs could be performed under good manufacturing practice conditions. Donor-reactive Tregs may be a valuable tool to control immune responses after transplantation a setting in which polyspecific Tregs have failed to date.

Traceability in fluorometry--Part I: Physical standards.

The inter-instrument, inter-laboratory, and long-term comparability of fluorescence data requires the correction of the measured emission and excitation spectra for the wavelength- and polarization-dependent spectral irradiance of the excitation channel at the sample position and the spectral responsivity of the emission channel employing procedures that guarantee traceability to the respective primary standards. In this respect the traceability chain of fluorometry is discussed from a radiometrist's point of view. This involves, in a first step, the realization of the spectral radiance scale, based on the blackbody radiator and electron storage ring, and the spectral responsivity scale, based on the cryogenic radiometer and their control via key comparisons of the national metrology institutes. In a second step, the characterization including state-of-the art uncertainties of the respective source and detector transfer standards such as tungsten strip lamps, integrating sphere radiators, and trap detectors used to disseminate these radiometric quantities to users of spectroscopic techniques is presented.