Altered placental macrophage numbers and subsets in pregnancies complicated with intrahepatic cholestasis of pregnancy (ICP) compared to healthy pregnancies.

INTRODUCTION: Intrahepatic cholestasis of pregnancy (ICP) can result in adverse outcomes for both mother and fetus. Inflammatory (M1 subset) or anti-inflammatory (M2 subset) macrophage polarisation is associated with various complications of pregnancy. However, the influence of ICP on macrophage numbers and polarisation remains unknown. This study analyses macrophage density and distribution in placentas of patients with ICP compared to controls. Clinical parameters were correlated to macrophage distribution and ursodeoxycholic acid use (UDCA). METHODS: This study included routinely collected placental tissue samples of 42 women diagnosed with ICP and of 50 control pregnancies. Immunohistochemical staining was performed on placental tissue using CD68 antibody as a pan-macrophage marker, CD206 antibody as an M2 and HLA-DR antibody as an M1 macrophage marker. Macrophage density (cells/mm2) and distribution (CD206+/CD68+ or CD206+/CD68+HLA-DR+) in both decidua (maternal tissue) and villous parenchyma (fetal tissue) were compared between groups. Macrophage density and distribution were correlated to clinical parameters for ICP patients. RESULTS: The density of CD68+ macrophages differed significantly between groups in villous parenchyma. In both decidua and villous parenchyma, CD206+/CD68+ ratio was significantly lower in ICP patients compared to controls (p = 0.003 and p=<0.001, respectively). No difference was found based on UDCA use or in CD68+HLA-DR+ cell density. Significant correlations were found between macrophage density and peak serum bile acids and liver enzymes. DISCUSSION: In ICP patients, an immune shift was observed in both decidual and villous tissue, indicated by a lower CD206+/CD68+ ratio. ICP seems to affect placental tissue, however more research is required to understand its consequences.

Alloimmunity and Cholestasis After Liver Transplantation in Children With Progressive Familial Intrahepatic Cholestasis.

OBJECTIVES: Bile salt export pump (BSEP) deficiency is an important reason for chronic cholestasis leading to liver transplantation (LT) in early childhood. The underlying pathology is a dysfunction of BSEP due to various mutations in the ABCB11 gene. Cases of clinical recurrence after LT due to alloantibodies directed against BSEP (antibody-induced BSEP deficiency [AIBD]) have been reported. Most of these patients could be controlled by intensified immunosuppression. METHODS: We here report on 3 children with BSEP-deficiency and end-stage liver disease, which developed AIBD after LT refractory to extensive immunosuppressive and immunomodulatory treatments; retransplantation was necessary in all 3 patients. In 1 patient, a stem cell transplantation was performed successfully. RESULTS: AIBD seems to be induced by triggering factors such as initial impaired graft function or infections after LT. CONCLUSIONS: The underlying mutation may play a role in this process. Intensifying immunosuppression may be able to control AIBD, but some cases seem to be refractory to treatment and require retransplantation. Stem cell transplantation may provide a new therapeutic option for cases refractory to conservative treatment.

Cholestasis After Pediatric Liver Transplantation-Recurrence of a Progressive Familial Intrahepatic Cholestasis Phenotype as a Rare Differential Diagnosis: A Case Report.

INTRODUCTION: Nonobstructive cholestasis after pediatric liver transplantation is a common diagnostic and therapeutic dilemma. We describe a girl with neonatal cholestasis because of progressive familial intrahepatic cholestasis 2 (PFIC-2) and presence of a homozygous splice site mutation in the ABCB11 gene. Liver transplantation was performed because of end-stage liver disease at the age of 6. Cholestasis with normal gamma-glutamyl transferase (GGT) developed 8 years after liver transplantation. A liver biopsy showed canalicular cholestasis and giant cell hepatitis without evidence of rejection, mimicking PFIC-2. Immunofluorescence staining of normal human liver sections with patient's serum revealed reactivity toward a canalicular epitope, which could be identified as bile salt export pump (BSEP) using BSEP-yellow fluorescent protein (YFP) transfected cells. Our patient developed a recurrence of a PFIC-2 phenotype due to production of antibodies against BSEP (alloimmune BSEP disease [AIBD]). Intensification of immunosuppressive therapy as well as antibody treatment with plasmapheresis and Rituximab were initiated, leading to stabilization of the clinical condition and depletion of anti-BSEP antibodies in serum. However, after 1 year liver transplantation was necessary again because of end-stage liver insufficiency. Afterward, immunomodulatory treatment consisted of tacrolimus, mycophenolate mofetil, prednisone, immunoadsorption, and high-dose immunoglobulin therapy (1 g/kg/d). CONCLUSION: Cholestasis after liver transplantation may indicate an AIBD with a PFIC-2 phenotype. Besides enhancement of immunosuppressive therapy, an antibody depletion with plasmapheresis, immunoadsorption, immunoglobulins, and B-cell depletion represents a therapeutic option.

Recovery from LCDD-associated Severe Liver Cholestasis: a Case Report and Literature Review.

Light chain deposition disease (LCDD) is a rare multisystemic disorder associated with plasma cell proliferation. It mainly affects the kidney, but liver and heart involvement may occur, sometimes mimicking the picture of systemic amyloidosis. Liver disease in LCDD is usually asymptomatic and exceptionally manifests with severe cholestatic hepatitis. We report the case of a 66-year-old female with κ-LCDD and cast nephropathy in the setting of symptomatic multiple myeloma who, after a first cycle of bortezomib-dexamethasone chemotherapy, developed severe and rapidly worsening intrahepatic cholestasis secondary to liver κ-light chain deposition. Intrahepatic cholestasis was attributed to LCDD on the basis of the liver histology and exclusion of possible diagnoses. Chemotherapy was maintained and resulted in progressive resolution of cholestasis. We report here an uncommon presentation of LCDD, with prominent liver involvement that fully recovered with bortezomib-based chemotherapy, and briefly review the relevant literature.

Role of macrophages in bile acid-induced inflammatory response of fetal lung during maternal cholestasis.

UNLABELLED: Infant respiratory distress syndrome (iRDS) in babies born from women with intrahepatic cholestasis of pregnancy (ICP) has been associated with intrauterine exposure to high bile acid levels. Here, we have investigated the role of macrophages in hypercholanemia-induced changes in maternal and fetal lung. Obstructive cholestasis in pregnant rats (OCP) was maintained from day 14 of gestation to term. Gene expression was determined by RT-QPCR, Western blot, and immunofluorescence. The maternal-fetal bile acid pool was radiolabelled using [(3)H]-taurocholate. OCP resulted in increased bile acids in maternal and fetal organs, including lungs. This was accompanied by structural changes in lung tissue, more marked in fetuses (peribronchial edema, collapse of alveolar spaces and deposits of hyaline material in the alveolar lumen), and infiltration of lung tissue by inflammatory cells. The abundance of macrophages and neutrophils in bronchoalveolar lavage fluid (BALF) was also increased in OCP group. Phospholipase A2-IIA (PLA2), the key enzyme in surfactant degradation, was mainly immunodetected in macrophages, which also expressed the bile acid receptor TGR5. The overall expression of PLA2 was markedly enhanced in maternal and fetal lungs of OCP group and in control maternal BALF cells incubated with bile acids. In neonates born from OCP mothers, the enhanced expression of erythropoietin suggested the presence of hypoxia due to iRDS. In conclusion, these results indicate that the accumulation of bile acids due to maternal cholestasis triggers an inflammatory response in the maternal and fetal lungs together with enhanced macrophage-associated PLA2 expression, which may play an important role in iRDS development. KEY MESSAGES: Maternal cholestasis causes respiratory distress syndrome in rat neonates. Cholestasis in pregnant rats causes bile acid accumulation in the fetal lung. This induces lung macrophages infiltration and inflammatory response. Alveolar macrophages co-express phospholipase A2-IIA and TGR5, but not FXR. Bile acid accumulation stimulates phospholipase A2-IIA, but not TGR5, expression.

Cholestatic liver disease after rituximab and adalimumab and the possible role of cross-reacting antibodies to Fab 2 fragments.

BACKGROUND: Millions of patients are treated with therapeutic monoclonal antibodies (Tmabs) for miscellaneous diseases. We investigated sera from six patients who received immune globulin, from one patient with refractory anti-neutrophil-cytoplasmic antibody (ANCA)-associated granulomatosis with polyangiitis (GPA) who developed two episodes of acute cholestatic liver disease, one after treatment with rituximab and a second after adalimumab and a healthy control group. METHODS: Three sera from the patient and six sera from patients who received immune globulin were analyzed for antibodies to rituximab and adalimumab by ELISA. Additionally, sera from the patients and from nine healthy blood donors were coated with the Fab fragment of an unrelated humanized monoclonal antibody, with human Fc proteins as well as a mouse IgG globulin. RESULTS: Viral serology for hepatitis A, B, C and autoantibodies specific for autoimmune liver disorders were negative. In all three sera from the patient antibodies to rituximab could be detected, but also antibodies to adalimumab were present even at time points when the patient had not yet received adalimumab, indicating cross reactivity between both substances. Testing against an unrelated human Fab fragment revealed positive results, indicating that the patient had antibodies against human Fab fragments in general. The Fc proteins were negative, and patients' sera did also not react with mouse IgG globulins. Remarkably, 2 out of 5 patients which were treated with immune globulin had antibodies against human Fab fragments in general whereas in none of the samples from healthy controls antibodies to Fab fragment could be detected. CONCLUSION: This is the first study demonstrating cholestatic liver disease induced by two different Tmabs. Cross - reacting antibodies to Fab2 fragments in general are probably involved. Further studies must show if these Fab2 antibodies in general are related with drug-induced side effects and accelerated drug clearance in patients on Tmab therapy.

Rituximab as therapy for the recurrence of bile salt export pump deficiency after liver transplantation.

Progressive familial intrahepatic cholestasis type 2 (PFIC2) results from recessive mutations in the adenosine triphosphate-binding cassette B11 gene, which encodes for bile salt export pump (BSEP). Liver transplantation (LT) is offered to PFIC2 patients with end-stage liver disease. Reports have described recurrent cholestasis in PFIC2 patients after transplantation, and this has been associated with immunoglobulin G antibodies to BSEP. High-titer anti-BSEP antibodies appear to correlate with episodes of cholestatic graft dysfunction. There is no established paradigm for treating antibody-mediated posttransplant BSEP disease. It appears to be refractory to changes in immunosuppressant medications that would typically be effective in treating allograft rejection. Taking what is known about its pathophysiology, we designed a treatment consisting of rituximab, a chimeric monoclonal anti-CD20 antibody, in combination with intravenous immunoglobulin and plasmapheresis. Using this approach, we report the successful management of 2 patients with antibody-mediated recurrence of PFIC2 after LT.

Effect of maternal cholestasis on TGR5 expression in human and rat placenta at term.

BACKGROUND & AIMS: TGR5 (Gpbar-1) is a plasma membrane-bound bile acid receptor expressed in several tissues, including liver, intestine and brain. High levels of TGR5 mRNA have been detected in human and rodent placenta, however, localization of the TGR5 protein has not been studied in this tissue. We aimed at characterizing TGR5 expression in placental tissue and investigated the effect of bile acids and progesterone metabolites, which accumulate during intrahepatic cholestasis of pregnancy (ICP), on receptor expression and localization. METHODS: TGR5 mRNA levels and cell-specific localization were determined by quantitative PCR and immunofluorescence, respectively. RESULTS: In human term placentas, TGR5 was mainly localized in fetal macrophages and to a lower extent in trophoblasts. In placentas from ICP patients and pregnant rats with obstructive cholestasis a marked down-regulation of TGR5 mRNA expression was observed. However, the cell-specific distribution of the TGR5 protein was unaffected. Besides bile acids, progesterone and its metabolites (5α-pregnan-3α-ol-20-one/5α-pregnan-3ß-ol-20-one), which increase in serum during ICP, were able to dose-dependently activate TGR5. In addition, progesterone metabolites but not their sulfated derivatives nor taurolithocholic acid, significantly down-regulated TGR5 mRNA and protein expression in isolated human macrophages and a macrophage-derived cell line. CONCLUSION: Since fetal macrophages and trophoblast cells are exposed to changes in the flux of compounds across the placental barrier, the expression of TGR5 in these cells together with its sensitivity to bile acids and progesterone metabolites regarding receptor activity and mRNA expression suggest that TGR5 may play a role in the effect of maternal cholestasis on the placenta.

[Expression of HLA-G protein in placental tissues and its influence on Th1/Th2 cytokines in peripheral blood in patients with intrahepatic cholestasis of pregnancy].

OBJECTIVE: To investigate the changes of human leukocyte antigen-G (HLA-G) protein expression and Th1/Th2 type cytokines in intrahepatic cholestasis of pregnancy (ICP) and their relativity to the etiology of ICP. METHODS: Peripheral blood and placental tissues were obtained from 26 ICP patients (the ICP group) and 22 normal pregnant women (the NP group) in the operation room for Cesarean birth. Immunohistochemistry was used to detect the expression of HLA-G protein in the placental tissues. Meanwhile we tested the concentrations of tumor necrosis factor-alpha (TNF-alpha) and interleukin-4 (IL-4) by enzyme-linked immunosorbent assay (ELISA) in the peripheral blood and checked the levels of TBA in the serum. RESULTS: TBA level in the ICP group was (27.05+/-6.08) micromol/L, significant higher than that in the NP group (4.35+/-2.68) micromol/L (P<0.01). The positive expression of HLA-G protein in extravillous trophoblast in the ICP group was significantly lower than that in the NP group (P<0.01). The mean optical density (MOD) of positive expression of HLA-G protein in the placenta tissues in the ICP group (52.91+/-7.19) was significantly lower than that in the NP group (69.26+/-7.72) (P<0.01). The concentration of TNF-alpha was significantly higher in the ICP group (101.31+/-19.30) pg/mL than that in the NP group (54.51+/-23.72) pg/mL (P<0.01). The concentration of IL-4 was lower in the ICP group (22.16+/-6.55) pg/mL than that in the NP group (31.69+/-8.25) pg/mL (P<0.01). The ratio of TNF-alpha/IL-4 was higher in the ICP group (4.52+/-1.91) than that in the NP group (1.72+/-0.61) (P<0.01). There was a negative correlation between the MOD of HLA-G protein and TNF-alpha (r=-0.98, P<0.01) in the ICP group. No correlation with IL-4 and TNF-alpha/IL-4 was seen (P>0.05). There was a positive correlation between TBA and TNF-alpha (r=0.99, P<0.01), and a negative correlation between TBA and the MOD of HLA-G protein (r=-1.00, P<0.01) in the ICP group. No correlation with IL-4 and TNF-alpha/IL-4 was seen (P>0.05). CONCLUSION: There is an imbalance of Th1/Th2 cytokines to the Th1 type in the peripheral blood of ICP patients. The expression of HLA-G protein in the placenta of ICP patients decreases, leading to an increase of Th1 type cytokines that may be one of the reasons for liver destroy in ICP.

De novo bile salt transporter antibodies as a possible cause of recurrent graft failure after liver transplantation: a novel mechanism of cholestasis.

Progressive familial intrahepatic cholestasis type 2 (PFIC-2) is caused by mutations of the bile salt export pump (BSEP [ABCB11]), an ATP-binding cassette (ABC)-transporter exclusively expressed at the canalicular membrane of hepatocytes. An absence of BSEP from the canalicular membrane causes cholestasis and leads to liver cirrhosis, which may necessitate liver transplantation in early childhood. We report on the first case of a child with PFIC-2 suffering from repeated posttransplant recurrence of progressive intrahepatic cholestasis due to autoantibodies against BSEP. These antibodies occurred after transplantation and were detected in the patient's serum and at the canalicular membrane of two consecutive liver transplants. The antibodies were reactive toward the first extracellular loop of BSEP, were of high affinity, and inhibited transport activity of BSEP, thus causing severe cholestasis. The patient had three homozygous, missense changes in the BSEP gene. Their combination resulted in the complete absence of BSEP, which explains the lack of tolerance, a prerequisite of autoantibody formation toward BSEP. The findings illustrate a novel disease mechanism due to a new class of functionally relevant autoantibodies resulting in cholestasis and subsequent liver failure.

Epidemiological, immunological and clinical characteristics of acute hepatitis C.

UNLABELLED: The aim of the study was to make a clinical and epidemiological and immunological characteristic of patients with acute hepatitis C infection (AHC). PATIENTS AND METHODS: The study included 178 patients with AHC; they were studied in terms of clinical course, biochemical constellations, T and B lymphocyte subpopulations, level of TNF-alpha in the blood serum, presence of autoantibodies, and the outcome of the disease in a five-year follow-up period. METHODS: anti-HCV (EIA), HCV-RNA (PCR), HCV genotyping; ALT, AST, AP, gamma-GT; ultrasonography and liver biopsy. RESULTS: AHC incidence increased six-fold between 2000 and 2006. The prevalence of the disease among intravenous drug-users (IDUs) was 46.07%. Young people (31.71 +/- 1.21) and males (67.98%) were prevalent. The genotype HCV-1 was prevalent. AHC ran with icterus in 70.22% of all cases, while it was anicteric in 29.78%; ALT-activity was high--it was mean 1007.94 +/- 59.87 U/l; intrahepatic cholestasis was found in 38.80%. A light form of the disease was found in 43.26%, mild--in 50.56%, and severe--in 6.18%, without reaching acute liver failure. In the acute stage of the disease, an increase of helper/inducer CD3+CD4+ (p = 0.001), memory T helper CD4+CD29+ (p < 0.0001), activated CD3+HLA-DR+ (p <0.0001), mature CD3+ T cells (p < 0.05), naive CD2+T (p < 0.01), and B-lymphocytes CD19+ (p < 0.001) was found, together with a non-significant increase of the suppressor/cytotoxic CD3+CD8+ T lymphocytes in comparison with the controls. The total killer CD56+ were reduced, as well as the MHC restricted killer cells CD8+CD56+. TNF-alpha was elevated in the serum in the light and mild forms (p < 0.0001). The participation of non-organ-specified antibodies (NOSAs) was minimal. Anti-MLA titer was 1/80 in two patients. Five years after the outset of AHC, a spontaneous viral clearance was established in 36.67% and chronic hepatitis in 63.33%. CONCLUSION: Despite the initially activated immune cellular response strongly correlating with a well expressed cytolytic syndrome around 2/3 of the AHC patients develop a chronic form of the disease.

[Detection of CD4+CD25+ regulatory T cells in liver tissues of fibrosing cholestatic hepatitis after liver and kidney transplantation].

OBJECTIVES: To study the expression and distribution of CD4+CD25+ regulatory T cells (Treg) in liver tissues of patients with fibrosing cholestatic hepatitis (FCH) after liver and kidney transplantation and to investigate their roles in the pathogenesis of FCH. METHODS: Liver biopsy specimens from five patients with FCH were studied histopathologically. A specific marker for CD4+CD25+ regulatory T cells in those specimens was detected with anti-FOXP3 monoclonal antibody by immunohistochemistry. Apoptoses of hepatocytes were detected with in situ apoptosis detection TUNEL kit. RESULTS: Fibrosis in portal and around portal areas, cholestasis in some of the hepatocytes and canaliculi, widespread ballooning and ground-glass appearance of liver cells, and positivity of HBsAg and HBcAg and Pre-S1 protein were seen in the livers of all cases. The positive signal of FOXP3 was located in the cytoplasm of lymphocytes and the positive cells were mainly aggregated in the portal areas as well as occasionally appearing in the hepatic sinusoids. There were many more apoptotic hepatocytes near the portal areas. CONCLUSION: Fibrosing cholestatic hepatitis has specific pathological characteristics which might be caused by high expressions of FOXP3 in liver tissues.

[The preliminary study on cord blood lymphocytes apoptosis in patients with intrahepatic cholestasis of pregnancy].

OBJECTIVE: Intrahepatic cholestasis of pregnancy (ICP) is a disease happening during pregnancy and does a great harm to the fetus. The etiology and pathogenesis of ICP are not clear. It is very important to investigate the relationship between ICP cord blood lymphocytes (CBLs) apoptosis and its clinical significance. METHODS: The CBLs were isolated from 20 patients with ICP and 20 normal pregnant women. Flow cytometry (FCM) was used to detect the apoptosis cells in the CBLs. The apoptosis regulatory molecules (Fas/FasL, Bcl-2) of CBLs were detected by immunohistochemical technique. RESULTS: The percentage of apoptotic CBLs in ICP patients were significantly lower than that in the normal pregnant women (P < 0.05). The FasL protein expression of CBLs in ICP patients was significantly higher than that in normal pregnant women (P < 0.05), but Fas and Bcl-2 protein expression were normal. CONCLUSION: The abnormal apoptosis of CBLs may influence the immune reaction of ICP patients, which does something in the pathogenesis of ICP.

Is a leaky gut involved in the pathogenesis of intrahepatic cholestasis of pregnancy?

Increased gastrointestinal permeability has been demonstrated in several liver diseases. It may facilitate the absorption of gut-derived endotoxin-stimulating Kupffer cells to release proinflammatory cytokines or other potentially hepatotoxic compounds. We examined gastrointestinal permeability, plasma levels of anti-lipopolysacharides (anti-LPS), and four proinflammatory cytokines in 20 patients with intrahepatic cholestasis of pregnancy (ICP) compared with 22 normal pregnant and 29 non-pregnant women. Urinary excretion of sucrose and the urinary lactulose/mannitol (L/M) ratio after a standard oral load were used to assess gastrointestinal permeability. Anti-LPS (IgA, IgM, and IgG) were measured in peripheral blood by Human EndoCAb test kit; TNF-alpha, IL-1beta, IL-6, and IL-10 by Quantikine HS human immunoassays. Sucrose urinary excretion was similar in the three groups, indicating normal gastric permeability. The urinary L/M ratio was significantly higher in ICP than in the other groups [median (interquartile range): 0.018% (0.011-0.023) in ICP, 0.012% (0.009-0.016) in normal pregnancies, and 0.009% (0.008-0.012) in non-pregnant women, P < .01]. No significant differences were found in anti-LPS or cytokines plasma levels except slightly higher levels of IL-6 in ICP patients than in non-pregnant women (P < .05). Four of five women with abnormal urinary L/M ratio during ICP continued to show abnormalities in tests up to 2 years after delivery. In conclusion, an increased intestinal permeability was detected in ICP patients during and after pregnancy. A "leaky gut" may participate in the pathogenesis of ICP by enhancing the absorption of bacterial endotoxin and the enterohepatic circulation of cholestatic metabolites of sex hormones and bile salts.

Acute liver failure in a renal transplant patient caused by adenoviral hepatitis superimposed on a fibrosing cholestatic hepatitis B.

We report the first case of fulminant liver failure due to necrotizing adenovirus hepatitis superimposed on a fibrosing cholestatic hepatitis B in an immunosuppressed patient after renal transplantation. Diagnosis was made in vivo by liver biopsy and confirmed at autopsy.

Centrilobular necrosis after orthotopic liver transplantation: association with acute cellular rejection and impact on outcome.

Several studies have linked centrilobular necrosis (CN) to acute cellular rejection (ACR) following liver transplantation. However, it may be difficult to establish the diagnosis of ACR when the classic portal features are absent. The aim of the present study was to identify specific features that would help to recognize ACR in biopsies with CN. One hundred and forty liver biopsies with CN were identified from 97 patients who underwent liver transplantation. The following histopathologic features were assessed: CN, steatosis, lobular inflammation, cholestasis, endothelialitis, and fibrosis. CN was graded semiquantitatively. A number of clinical and biochemical parameters were also recorded. Biopsies with CN were assessed for the presence or absence of ACR and divided into two groups accordingly. The associations of the biochemical, pathologic, and clinical features with ACR were assessed using a multivariate logistic regression model. The outcomes of patients with and without rejection were compared using the Cox proportional hazards regression model. Seventy-four biopsies (52.9%) showed evidence of ACR, and 52 patients (53.6%) had evidence of ACR at the first biopsy with CN. The multivariate analysis showed the presence of cholestasis, lobular inflammation, the ALT level, and time since liver transplantation to be independent predictors of the presence of ACR in biopsies with CN. Patients with ACR on their first biopsy with CN were significantly more likely to experience graft loss compared with patients without ACR. In conclusion, the presence of cholestasis and lobular inflammation on biopsies with CN appeared helpful in predicting its association with ACR.

Effects of proinflammatory cytokines on rat organic anion transporters during toxic liver injury and cholestasis.

Hepatobiliary transporters are down-regulated in toxic and cholestatic liver injury. Cytokines such as tumor necrosis factor alpha (TNF-alpha) and interleukin 1 beta (IL-1 beta) are attributed to mediate this regulation, but their particular contribution in vivo is still unknown. Thus, we studied the molecular mechanisms by which Ntcp, Oatp1, Oatp2, and Mrp2 are regulated by proinflammatory cytokines during liver injury. Rats were injected intraperitoneally with either carbon tetrachloride or endotoxin. Inactivation of TNF-alpha and IL-1 beta was achieved by repetitive intraperitoneal injection of etanercept and anakinra, respectively. Messenger RNA (mRNA) levels of transporters and binding activities as well as nuclear protein levels of Ntcp, Oatp2, and Mrp2 transactivators were determined 20 to 24 hours later. In contrast to IL-1 beta, TNF-alpha inactivation alone fully prevented down-regulation of Ntcp, Oatp1, and Oatp2 mRNA as well as reduced binding activity of hepatocyte nuclear factor 1 (HNF-1) in CCl(4)-induced toxic injury. In endotoxemia, down-regulation of Mrp2, and partially in case of Ntcp, could be prevented by IL-1 beta but not TNF-alpha blockade. However, inactivation of either cytokine led to preservation of HNF1 and partially of retinoid X receptor/retinoic acid receptor (RXR/RAR) binding activity. No effect of anticytokines was seen on pregnane X receptor (PXR) and constitutive androstane receptor (CAR) binding activity as well as nuclear protein mass. In conclusion, TNF-alpha represents the master cytokine responsible for HNF1-dependent down-regulation of Ntcp, Oatp1, and Oatp2 in CCl(4)-induced toxic liver injury. IL-1 beta predominates in a complex signaling network of Ntcp and Mrp2 regulation in cholestatic liver injury. In contrast to in vitro studies, HNF1 and RXR/RAR-independent mechanisms appear to be more important in regulation of Mrp2 and Ntcp gene expression in endotoxemia.

Allergic cholestatic hepatitis and exanthema induced by metamizole: verification by lymphocyte transformation test.

We report about a 66-year-old-male patient who was hospitalized with generalized exanthema and increase of liver enzymes after intake of metamizole because of flue-like symptoms. Despite initial high dose steroids, disease activity persisted, and therefore liver biopsy was performed. Histology revealed acute hepatitis with perivenular non-bridging confluent necrosis and granuloma formation consistent with drug-induced hepatitis. A metamizole-induced process was suspected. Lymphocyte transformation test confirmed the sensitization of the patient's lymphocytes to metamizole and three of its four metabolites (4-methylaminoantipyrine, 4-acetylaminoantipyrine and 4-formylaminoantipyrine). Other drugs could be excluded with high probability. In the follow-up, the general condition of the patient improved, and liver enzymes decreased under treatment with steroids. Thus, we conclude that in this patient metamizole has induced an allergic reaction not only of the skin but also of the liver. To our knowledge, an allergic cholestatic hepatitis caused by metamizole has been reported only once in literature.

Amelioration of steroid-resistant chronic graft-versus-host-mediated liver disease via tacrolimus treatment.

Liver disease associated with chronic graft-versus-host disease (cGVHD) is a major cause of mortality and morbidity. Steroids and cyclosporin (CSA), which are the standard therapy, give rather disappointing results, and toxicity is high. Tacrolimus (FK506) is a potent macrolide lactone immunosuppressant that is used in the prevention of solid organ rejection. This study evaluated the therapeutic role of FK506 in the treatment of severe cGVHD-mediated liver disease that did not respond to combined steroids and CSA therapy. Fifteen patients with various hematological disorders who underwent allogeneic stem cell transplantation were enrolled in the study. All patients had severe cholestatic liver disease disturbances and underwent liver biopsy, which was compatible with cGVHD-mediated liver disease. All the patients were negative for markers of chronic liver disease, including viral serology. They received FK506 orally (4-20 mg/day according to serum levels), and were evaluated biweekly by physical examination and liver function tests. Patients were followed for a median of 12 months (range 3-24 months). FK506 treatment ameliorated liver functions in 9 of 15 patients (60%), 5 of whom demonstrated complete normalization of liver enzymes (33%). In 5 patients, no major effect was observed, and 1 patient showed deterioration of his liver functions. Mean GGT levels decreased from 171.5 to 55.6 within 6 months of treatment. Median time to response was 3 months (range 1-11). Side effects were generally transient. Treatment with FK506 was found to be effective in the majority of patients with steroid and CSA-resistant cGVHD-associated liver disease, with manageable side effects. In view of these findings, FK506 may yet evolve into first line therapy for cGVHD induced liver toxicity.