Sabine Boas-Knoop

Relationship between the interleukin‐28b gene polymorphism and the histological severity of hepatitis C virus–induced graft inflammation and the response to antiviral therapy after liver transplantation

Up to 30% of liver transplants will develop graft cirrhosis within 5 years after liver transplantation (LT) due to recurrent HCV‐infection forwarding accelerated graft damage. Genetic variants of cytokines involved in the immune response may contribute to the degree of graft inflammation, fibrosis progression, and antiviral therapy outcome. The aim of our study was to analyze biochemical and histological inflammation extent based on protocol liver biopsies and to evaluate the role of genetic variants of IL‐28b in HCV‐related graft disease and antiviral treatment response. 183 patients, who underwent liver transplantation for HCV‐induced liver disease, were genotyped for IL‐28b (rs8099917, G ≥ T) by TaqMan Genotyping Assay. 56 of 159 patients have been successfully treated with interferon‐based antiviral therapy. 605 protocol liver biopsies performed 0.5 to 10 and more than 10 years after transplantation were evaluated according to Desmet and Scheuer classification of inflammation and fibrosis. Prevalence of IL‐28b‐genotypes was correlated with histological severity of graft damage, levels of aminotransferases, occurrence of acute cellular rejection, pre‐treatment viremia, and antiviral therapy outcome. Significant association of IL‐28b‐genotype distribution was observed to the median grade of inflammation (p < 0.001), mean levels of aminotransferases (ALT: p = 0.001, AST: p = 0.003), median pre‐treatment viremia level within 1 year after LT (p = 0.046) and interferon‐based antiviral therapy failure (p < 0.001). Among successfully treated patients, G‐allele was significantly less frequent, and the genotype GG was not present at all. No differences were observed regarding acute cellular rejection (p = 0.798) and fibrosis stages (p = 0.586). IL‐28b polymorphism seems to influence the degree of graft inflammation at biochemical and histological levels. G‐allele might serve as a marker for graft inflammation and as a predictor for unfavorable antiviral therapy outcome in HCV‐re‐infected LT‐population. Liver Transpl, 2011.

Erythropoietin reduces ischemia-reperfusion injury in the rat liver.

Background: Human recombinant erythropoietin (Epo) has recently been shown to be a potent protector of ischemic damage in various organ systems. A significant reduction of stroke injury following cerebral ischemia has been postulated as well as improved cardiomyocyte function after myocardial infarction in tissue pretreated with Epo. It was the aim of this study to evaluate the effects of Epo in liver ischemia. Material and Methods: Rats were subjected to 45 min of warm hepatic ischemia. Animals were either pretreated with 1,000 IU of Epo in three doses or received 1,000 IU into the portal vein 30 min before ischemia. Control animals received saline at the same time points before ischemia. Animals were than sacrificed 6, 12, 24, 48 h and 7 days after surgery and transaminases were measured. Liver specimens were evaluated regarding apoptosis, necrosis and regeneration capacity. Results: Apoptosis rates were dramatically reduced in animals pretreated with Epo while mRNA of tumor necrosis factor-α and STAT-3 were upregulated in all groups. Intraportal venous injection displayed superiority to subcutaneous preconditioning. Transaminases were significantly reduced among the Epo-treated animals after 6 and 12 h. Conclusion: Our data suggests a protective effect of Epo in warm hepatic ischemia and reperfusion injury in the rat.

Erythropoietin promotes hepatic regeneration after extended liver resection in rats

Background and Aim:  It has been proven in various animal studies that recombinant human erythropoietin (rHuEPO) protects renal, cardiac and neuronal, as well as hepatic, tissue from ischemia, and promotes regeneration of adult central nervous system neurons. To date, no data are available as to whether rHuEPO has the ability to stimulate liver regeneration after liver resection.

Transforming Growth Factor β1 Polymorphisms and Progression of Graft Fibrosis After Liver Transplantation for Hepatitis C Virus-Induced Liver Disease

Re‐infection with the hepatitis C virus (HCV) is an important development after liver transplantation (LT); it can lead to graft fibrosis. The aim of this study was to assess the role of transforming growth factor β1 (TGF‐β1) polymorphisms in the development of HCV‐related graft disease by evaluating protocol liver biopsies. A total of 192 patients with a recurrence of HCV infection after LT were genotyped for TGF‐β1 codon 10 (C→T) and codon 25 (G→C) using the polymerase chain reaction. Histological evaluation of 614 protocol liver biopsies obtained from these patients was undertaken using the classification of Desmet and Scheuer to stage the degree of fibrosis. Mild stages of fibrosis (0‐2) were compared to advanced stages of fibrosis (3‐4) that developed during the period of infection with the virus. Correlations between the prevalence of TGF‐β1 genotypes and the different degrees of fibrosis that developed were determined. No statistically significant differences were found for genotype distributions (codons 10 and 25) with respect to recipient age, donor sex, occurrence of acute cellular rejection, and response to antiviral therapy. However, the C allele at codon 25 was significantly less frequent in the group with advanced fibrosis (P = 0.001). Furthermore, a positive association was found between progression of fibrosis and male recipient sex (P = 0.024), donor age (P = 0.041), and viral genotype 1b (P = 0.002). In conclusion, this study, in which the evolution of hepatic fibrosis was assessed histologically in a large cohort of patients with HCV re‐infection after LT, has demonstrated that the C allele at codon 25 of the TGF‐β1 gene is a marker for the development of graft fibrosis. Liver Transpl, 2011.

Erythropoietin reduces ischemia‐reperfusion injury after liver transplantation in rats

Human recombinant Erythropoietin (rHuEpo) has recently been shown to be a potent protector of ischemia‐ reperfusion injury in warm‐liver ischemia. Significant enhancement of hepatic regeneration and survival after large volume partial hepatic resection has also been demonstrated. It was the aim of this study to evaluate the capacities of rHuEpo in the setting of rat liver transplantation. One‐hundred‐and‐twenty Wistar rats were used: 60 recipients received liver transplantation following donor organ treatment (60 donors) with either 1000 IU rHuEpo or saline injection (controls) into portal veins (cold ischemia 18 h, University of Wisconsin (UW) solution). Recipients were allocated to two groups, which either received 1000 IU rHuEpo at reperfusion or an equal amount of saline (control). Animals were sacrificed at defined time‐points (2, 4.5, 24, 48 h and 7 days postoperatively) for analysis of liver enzymes, histology [hematoxylin‐eosin (HE) staining, periodic acid Schiff staining (PAS)], immunostaining [terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), Hypoxyprobe] and real‐time polymerase chain reaction (RT‐PCR) of cytokine mRNA (IL‐1, IL‐6). Lactate dehydrogenase (LDH) and alanine aminotransferase (ALT) values were significantly reduced among the epo‐treated animals 24 and 48 h after liver transplantation (LT). The TUNEL and Hypoxyprobe analyses as well as necrotic index evaluation displayed significant reduction of apoptosis and necrosis in rHuEpo‐treated graft livers. Erythropoietin reduces ischemia‐reperfusion injury after orthotopic liver transplantation in rats.

Hedgehog pathway as a potential treatment target in human cholangiocarcinoma

Innovative treatment concepts targeting essential signaling pathways may offer new chances for patients suffering from cholangiocarcinoma (CCC). For that, we performed a systematic molecular genetic analysis concerning the Hedgehog activity in human CCC samples and analyzed the effect of Hh inhibition on CCC cells in vitro and in vivo.

Improvement of Microsurgical Techniques in Orthotopic Rat Liver Transplantation

BACKGROUND Because of the limited tolerance to portal venous clamping, the model of liver transplantation in rats represents a difficult task, which requires a great proportion of experience. Since techniques that include the introduction of an artificial stent increase the risk of thrombosis, it was our goal to modify the classical vascular hand-sewn venous anastomosis technique by using a modified end-to-end knotless procedure. MATERIALS AND METHODS Seventy-two animals were randomly assigned to 1 of 2 experimental groups, which differed by the technique for the 3 venous anastomoses (supra- and infrahepatic vena cava, portal vein). Group 1 comprised the established suturing technique for rat liver transplantation, whereas all venous anastomosis of the second group were performed using our modified technique. RESULTS With our method, average anhepatic time could be significantly reduced from 14 min 10 s (+/-100 s) to 11 min 40 s (+/-60 s) (P < 0.001). Kaplan-Meier survival rates demonstrated a better 7-d survival for the knotless (94%) compared to the classic technique (83%) (not significant, P = 0.137). Biliary complications were low in both groups but tended to be higher in the classical group. CONCLUSIONS Our modified knotless anastomosis proves to be equally safe in regard to complications, improves timing, and provides excellent results in the model of orthotopic rat liver transplantation.

Association of mannose-binding lectin-2 gene polymorphism with the development of hepatitis C-induced hepatocellular carcinoma.

Background: Development of end‐stage liver and graft disease is suspected to be partially determined by the individual genetic background. Mannose‐binding lectin (MBL) is an important immunomodulatory factor, which is supposed to be involved in complement activation and oncogenesis. Genetic polymorphisms of MBL‐2 alter MBL functionality. The aim of our study was to determine the prevalence of MBL‐2 polymorphism (rs7096206) in hepatitis C virus (HCV)‐induced hepatocellular carcinoma (HCC) based on histological analysis of explanted livers in patients undergoing liver transplantation (LT).

Role of mannose‐binding lectin‐2 polymorphism in the development of acute cellular rejection after transplantation for hepatitis C virus‐induced liver disease

The development of liver and graft disease is suspected to be affected by genetic diversity. Mannose‐binding lectin‐2 (MBL‐2) is an important immunomodulatory factor that is involved in complement activation. The aim of our study was to elucidate the role of MBL‐2 genotypes after liver transplantation (LT) for hepatitis C virus (HCV)‐induced liver disease regarding the incidence of acute cellular rejection (ACR), graft inflammation, fibrosis development, and antiviral treatment response.

CD44 and CXCL9 serum protein levels predict the risk of clinically significant allograft rejection after liver transplantation

The diagnosis of acute cellular rejection (ACR) after liver transplantation is based on histological analysis of biopsies because noninvasive biomarkers for allograft rejection are not yet established for clinical routines. CD31, CD44, and chemokine (C‐X‐C motif) ligand (CXCL) 9 have previously been described as biomarkers for cross‐organ allograft rejection. Here, we assessed the predictive and diagnostic value of these proteins as serum biomarkers for clinically significant ACR in the first 6 months after liver transplantation in a prospective study. The protein levels were measured in 94 patients immediately before transplantation, at postoperative days (PODs) 1, 3, 7, and 14 and when biopsies were performed during episodes of biochemical graft dysfunction. The CD44 serum protein levels were significantly lower at POD 1 in patients who experienced histologically proven ACR in the follow‐up compared with patients without ACR (P < 0.001). CXCL9 was significantly higher before transplantation (P = 0.049) and at POD 1 (P < 0.001) in these patients. Low CD44 values (cutoff, <200.5 ng/mL) or high CXCL9 values (cutoff, >2.7 ng/mL) at POD 1 differentiated between rejection and no rejection with a sensitivity of 88% or 60% and a specificity of 61% or 79%, respectively. The combination of both biomarker cutoffs at POD 1 had a positive predictive value of 91% and a negative predictive value of 67% for clinically significant ACR. Moreover, CD44 was significantly lower at the time of ACR (P < 0.001) and differentiated the rejection group from patients with graft dysfunction due to other reasons. Our results suggest that CD44 and CXCL9 may serve as predictive biomarkers to identify liver allograft recipients at risk for clinically significant ACR. Liver Transpl 21:1195–1207, 2015.