Non-viral Cirrhosis Research Articles

Model for End-stage Liver Disease and Model for End-stage Liver Disease-Na Scores Predict Both Before-Listing and Wait-List Mortality

BackgroundDue to the organ shortage, many patients die without transplantation, even before completing an evaluation for candidacy. We analyzed outcomes after patient referral and factors associated with mortality both before and after listing for cadaveric donor liver transplantation. MethodsWe analyzed 132 consecutive patients who were evaluated for candidacy for cadaveric donor liver transplantation between 2003 and 2010. ResultsThe study included 69 men and 63 women of median age 49 years (range, 1–65). Etiologies of diseases were acute hepatic failure (n = 19), liver cirrhosis due to hepatitis B or C (n = 36), primary biliary cirrhosis (n = 19), nonviral cirrhosis (n = 14), hepatocellular carcinoma (n = 13), or other causes (n = 31). After evaluation for candidacy, we listed 68 (52%), subjects whereas 24 (18%) died before listing. Factors affecting death before listing were the levels of albumin (P < .001), bilirubin (P < .001), sodium (P < .001), international normalized ratio (INR; P < .001), Model for End-stage Liver Disease (MELD) score (P < .001), MELD-Na score (P < .001), and Child-Pugh-Turcotte (CPT) score (P < .001). Based on multivariate Cox regression analysis, MELD score (hazard ratio [HR] 1.201, P = .017), MELD-Na score (HR 1.244, P = .014), CPT score (HR 1.468, P = .033), and INR (HR 0.491, P = .027) were independently associated with death before listing. Among 68 listed candidates, 11 (16%) underwent transplantation, whereas 29 (43%) died without transplantation. Based on multivariate Cox regression analysis, MELD score (HR 1.102, P = .001), MELD-Na score (HR 1.128, P = .001), and CPT score (HR 1.282, P = .038) independently predicted wait-list mortality. All 11 patients who underwent cadaveric liver transplantation were alive at 29 months (range, 1–55) after transplantation. ConclusionsPatients with a higher MELD, higher MELD-Na, and higher CPT score at referral were at greater risk for death without transplantation, especially before listing. Evaluation for transplantation candidacy is a time-consuming process. Therefore, earlier referral is mandatory to achieve successful listing for transplantation.

Association of Biochemical, Cytokine and Echocardiographic Markers of Cardiovascular Injuries with T894G Polymorphism of Endothelial Nitric Oxide Synthase Gene in Patients with Nonviral Liver Cirrhosis

Association of Biochemical, Cytokine and Echocardiographic Markers of Cardiovascular Injuries with T894G Polymorphism of Endothelial Nitric Oxide Synthase Gene in Patients with Nonviral Liver Cirrhosis

IL-28B rs12979860 C/T allele distribution in patients with liver cirrhosis: Role in the course of chronic viral hepatitis and the development of HCC

A single nucleotide polymorphism (rs12979860 C/T) 3kb upstream of the interleukin 28B (IL-28B) gene was shown to be associated with hepatitis C clearance. We verified whether this association also translates into a different genotype distribution at the end of the disease trajectory. A RFLP-PCR technique was used to genotype 412 patients with cirrhosis due to hepatitis C (n=199), hepatitis B (n=75), alcohol (n=110), and other causes (n=28), of whom 256 underwent liver transplantation (OLT). Hepatocellular carcinoma (HCC) was demonstrated in the native liver of 85 OLT patients, 52 with viral cirrhosis, and 33 with non-viral cirrhosis respectively. A group of 292 patients (235 HCV and 57 HBV positive) with mild chronic hepatitis and 344 healthy subjects served as controls. A significant difference (p=0.0005) was observed in IL-28B rs12979860 genotype frequencies between patients with viral cirrhosis (C/C=99, C/T=137, T/T=38) and those with non-viral cirrhosis (C/C=72, C/T=58, T/T=8). Patients with HCV related cirrhosis carried more frequently the T/T genotype in comparison to mild hepatitis C or HBV-related cirrhosis. IL-28B rs12979860 genotype frequencies were C/C=23, C/T=50, T/T=12 among OLT patients with cirrhosis complicated by HCC, and C/C=79, C/T=78, T/T=14 among patients with cirrhosis not complicated by HCC (p<0.005). IL-28B rs12979860 C/T polymorphism T allele is more prevalent in patients with viral cirrhosis due to HCV in comparison to other aetiologies and to patients with mild chronic hepatitis C. Among OLT patients, carriage of this allele seems to augment the risk of developing HCC.

Chapter 2: Diagnosis and surveillance

Chapter 2: Diagnosis and surveillance

Bax Inhibitor-1 down-regulation in the progression of chronic liver diseases

BackgroundBax inhibitor-1 (BI-1) is an evolutionary conserved endoplasmic reticulum protein that, when overexpressed in mammalian cells, suppresses the apoptosis induced by Bax, a pro-apoptotic member of the Bcl-2 family. The aims of this study were: (1) to clarify the role of intrinsic anti- and pro-apoptotic mediators, evaluating Bax and BI-1 mRNA and protein expressions in liver tissues from patients with different degrees of liver damage; (2) to determine whether HCV and HBV infections modulate said expression.MethodsWe examined 62 patients: 39 with chronic hepatitis (CH) (31 HCV-related and 8 HBV-related); 7 with cirrhosis (6 HCV-related and 1 HBV-related); 13 with hepatocellular carcinoma (HCC) [7 in viral cirrhosis (6 HCV- and 1 HBV-related), 6 in non-viral cirrhosis]; and 3 controls. Bax and BI-1 mRNAs were quantified by real-time PCR, and BI-1 protein expression by Western blot.ResultsCH tissues expressed significantly higher BI-1 mRNA levels than cirrhotic tissues surrounding HCC (P < 0.0001) or HCC (P < 0.0001). Significantly higher Bax transcripts were observed in HCV-genotype-1-related than in HCV-genotype-3-related CH (P = 0.033). A positive correlation emerged between BI-1 and Bax transcripts in CH tissues, even when HCV-related CH and HCV-genotype-1-related CH were considered alone (P = 0.0007, P = 0.0005 and P = 0.0017, respectively).ConclusionsBI-1 expression is down-regulated as liver damage progresses. The high BI-1 mRNAs levels observed in early liver disease may protect virus-infected cells against apoptosis, while their progressive downregulation may facilitate hepatocellular carcinogenesis. HCV genotype seems to have a relevant role in Bax transcript expression.

Adult to adult living-related liver transplant: Report on an initial experience in Italy

Introduction Living-related liver transplantation has become the treatment of choice for many liver diseases. We present our initial analysis of 53 cases of adult to adult living-related liver transplantation performed in a single institute in Italy. Materials and methods From January 2002 to September 2006, we performed 53 adult to adult living-related liver transplantations. The donors (age 18–53) all had genetic or emotional relationships; they were all ABO identical or compatible. Recipients (ages 18–68) suffered from cirrhosis secondary to viral etiology (18), hepatocellular carcinoma with viral cirrhosis (24), cystic fibrosis (2), primary biliary cirrhosis (2), hepatocellular carcinoma with non-viral cirrhosis (2), alcoholic cirrhosis (1), ornithine transcarbamylase deficiency (OTC), (1) criptogenic cryptogenic cirrhosis, (1) primary sclerosing cholangitis, (1) biliary atresia and metastatic carcinoid (1). Donor liver resection resulted in 51 right hepatectomies and two left hepatectomies. Graft body weight ratio was always above 0.8%; graft implantation was performed with the piggy back technique and, in 43 cases, with the use of veno-venous bypass. Results There was neither donor mortality nor need of blood transfusion. Actuarial recipient survival rate at 3 years was 82.66% and graft survival rate was 75.34%. Six patients underwent retransplantation: in four cases due to hepatic artery thrombosis, and in two, due to graft dysfunction. Three patients had one episode each of acute cellular rejection. Conclusion Adult to adult living-related liver transplantation represents a resource to be used in confronting organ shortage, and is a valuable option for decreasing mortality and drop out from the waiting list.

High prevalence of fatigue in chronic hepatitis and hepatitis C based cirrhosis in contrast to non-viral cirrhosis

High prevalence of fatigue in chronic hepatitis and hepatitis C based cirrhosis in contrast to non-viral cirrhosis

L/I‐2 Living related liver transplants: preliminary report on an initial experience in Italy

Introduction: Living related liver transplantation has become the treatment for many liver diseases. We present our initial analysis on 42 cases of adult‐to‐adult living related liver transplants (ALRLTs) performed in a single institution in Italy.Materials and methods: From January 2002 to February 2006, we performed 42 ALRLTs. The donors (age 18 to 53) all had genetic or emotional relationships; they were all ABO‐identical or ABO‐compatible. Recipients (age 18 to 68) suffered from cirrhosis secondary to viral etiology (n=13), hepatocellular carcinoma on viral cirrhosis (n=21), cystic fibrosis (n=2), primary biliary cirrhosis (n=2), hepatocellular carcinoma on nonviral cirrhosis (n=2), alcoholic cirrhosis (n=1), and metastatic carcinoid to the liver (n=1). Donor liver resection resulted in 40 right hepatectomies and 2 left hepatectomies. Graft body weight ratio was always above 1%; graft implantation was performed with the piggyback technique, and in 32 cases with the use of venovenous bypass.Results: There was no donor mortality; 4 donors had biliary complications. The actuarial recipient survival rate at 3 years was 89.25% and the graft survival rate was 82.69%. Graft loss occurred in 7 patients: 5 patients underwent retransplants (4 because of hepatic artery thrombosis, 1 because of graft dysfunction), and 2 patients had 1 episode of acute cellular rejection.Conclusion: ALRLTs represent a resource to face the organ shortage in a region where the deceased donor donation rate is 7.2 donors per year per million.

Brief Communication: Glomerulonephritis in Patients with Hepatitis C Cirrhosis Undergoing Liver Transplantation

Patients infected with hepatitis C virus (HCV) frequently develop renal failure after liver transplantation. To describe renal histologic characteristics and concomitant clinical features in HCV-infected patients with end-stage cirrhosis. Case series. Single-center liver transplant program in the United States. 30 patients who received liver transplants for HCV-induced cirrhosis. Kidney biopsy during liver engraftment. Clinical data and laboratory tests of renal function within 6 months before liver transplantation. Twenty-five patients had immune-complex glomerulonephritis: membranoproliferative glomerulonephritis type 1 (n = 12), IgA nephropathy (n = 7), and mesangial glomerulonephritis (n = 6). Of these patients, 10 had normal serum creatinine levels, normal urinalysis results, and normal quantitative proteinuria. For 5 others, the only renal abnormality was an increased serum creatinine level. No patient had cryoglobulins in the blood or kidney. This small observational study did not include patients with nonviral cirrhosis and did not document post-transplantation outcomes. Immune-complex glomerulonephritis was common in patients with end-stage HCV-induced cirrhosis and was often clinically silent. Its potential to cause renal failure after liver transplantation may be underappreciated.

Evidence for non-HFE linked hemochromatosis in Asian Indians

Hereditary hemochromatosis is commonly due to two HFE1 (Histone Family E1) gene mutations - H63D and C282Y. Mutations in the Asian Indians are less well studied. The aim of this preliminary study was to find out the prevalence of HFE gene mutations in nonviral liver cirrhosis patients. Unexplained liver cirrhosis cases with transferrin saturation> 45%, attending the gastroenterology clinic in the years 2004 and 2005 were subjects of the prospective study. Asymptomatic individuals with negative family history of hemolytic anemia or liver disease served as controls. The clinical presentation was recorded in the patients. Transferrin saturation was estimated by standard colorimetric technique. The two common mutations in HFE1 gene and Y250X mutation of TFR (transferrin receptor) gene were studied by polymerase chain reaction based methods. A majority of the cases were sporadic, but family history was positive in four patients. In one family with multiple affected members, there was clear evidence of autosomal dominant inheritance. Seven out of 31 (22.6%) of unexplained cirrhosis cases were positive for mutations. One was homozygous for H63D. In healthy controls, prevalence was 8.1% (6/74). None of the patients or controls was positive for C282Y mutation of HFE1 or Y250X of TFR gene. Thus, in a number of cases of hemochromatosis in Indians, a gene with dominant inheritance may be involved in causation of the phenotype. The prevalence of HFE mutations in Indians is comparable to that reported from neighboring countries. It is worth studying other mutations in HFE gene and other iron overload genes in cryptogenic cirrhosis cases.

Increased Populations of Regulatory T Cells in Peripheral Blood of Patients with Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide with a poor prognosis and one for which immunotherapy remains a viable option. Experimental tumor models have shown that regulatory T cells, a functionally unique subset of T cells, can suppress effective antitumor immune responses. This suppression might explain the poor outcome of some of the immunotherapy protocols currently being used. A better understanding of the role of regulatory T cells in HCC is important for design of future immunotherapy-based clinical protocols. We have studied regulatory T cells from 84 patients with HCC and 74 controls, including healthy donors, patients with chronic hepatitis B virus and hepatitis C virus infection and nonviral liver cirrhosis. Regulatory T cells were identified by fluorescence-activated cell sorting using a panel of antibodies and by real-time PCR analysis for Foxp3 expression. Functional studies were done to analyze their inhibitory role. Finally, regulatory T cells were analyzed in tumors and ascites from patients with HCC. Patients with HCC have increased numbers of CD4+CD25+ regulatory T cells in their peripheral blood, which express high levels of HLA-DR, GITR, and low or no CD45RA. These cells were anergic toward T-cell receptor stimulation and, when cocultured with activated CD4+CD25- cells, potently suppressed their proliferation and cytokine secretion. There were also high numbers of regulatory T cells in tumor-infiltrating lymphocytes of HCC patients comparable with the increase in their peripheral blood. Our data suggest that the increase in frequency of regulatory T cells might play a role in modulation of the immune response against HCC and could be important in design of immunotherapeutic approaches.

Cryopreserved human hepatocytes from cell bank: In vitro function and clinical application

We Aimed to analyze the in vitro function of isolated and cryopreserved human hepatocytes (CHH) from a cell bank and to define their potential clinical application in a bioartificial liver (BAL) device. Methods Over 24 months, 103 not transplantable livers were utilized for human hepatocytes isolation and cryopreservation. Hepatocytes isolated by collagenase were analyzed for yield, viability, diazepam metabolism, and production of human albumin after isolation and cryopreservation in LN 2. Results The causes for refusal for transplantation were macrosteatosis >60%, ischemic damage due to donor hypotension, and nonviral cirrhosis in 60%, 11%, and 8%, respectively. Cell yields averaged 7 million hepatocytes per gram of liver of mean viability of 80% ± 13%. The viability of CHH after thawing averaged 50%. Thawed hepatocytes showed diazepam metabolism, and human albumin synthesis comparable to fresh cells. CHH were utilized as the biological component of a BAL for temporary support as three applications of two patients affected by fulminant hepatic failure awaiting urgent transplant. Ten to 13 billion viable CHH were loaded into each BAL. Liver function showed bilirubin and ammonia reduction at the end of each treatment. One patient was successfully bridged to emergency OLTx after one BAL; in the second case there was spontaneous recovery of liver function after two BAL. Conclusions Recovery of donor human livers unwanted for transplantation allowed isolation and cryopreservation of viable and functionally active human hepatocytes, which have been banked and successfully used for clinical applications of a BAL device.

CRYOPRESERVED HUMAN HEPATOCYTES FROM CELL BANK: IN VITRO FUNCTION AND CLINICAL APPLICATION

P658 Aims: the development of a National Bank of Human Hepatocytes, isolated from livers unwanted for transplantation at harvesting, allows to obtain live hepatic tissue usable for in vitro and clinical evaluation of cell therapy and transplantation and bioartificial organs. The aim of this study is to analyze in vitro function of isolated and cryopreserved human hepatocytes from cell Bank and define their potential clinical application in a bioartificial liver (BAL) device. Methods: 103 not transplantable livers were utilized for human hepatocytes isolation and cryopreservation from July 2001 to December 2003 in the Nord Italia Transplant area (NITp and AIRT). Hepatocytes were isolated by collagenase-enzymatic digestion. Isolated hepatic cells were analyzed in terms of yield, viability (tripan blue and XTT test), diazepam metabolism (cyt P450 activity) and production of human albumin (immunonephelometry and mRNA by RT-PCR) after isolation (fresh hepatocytes) and after cryopreservation (10% DMSO) in liquid nitrogen. Results: macrosteatosis >60%, ischemic damage due to prolonged donor hypotension or cardiac arrest and non-viral cirrhosis were the causes of refusal of the liver for transplantation respectively in 60%, 11% and 8% of cases. Cell yield averaged 7 million of hepatocytes per gram of liver tissue digested; mean viability was 80%±13%. The best results in terms of cell yield and viability were obtained from steatotic organs, while cirrhotic livers yielded less cell/gr of liver (3x106 vs 7x105 p<0.043 and 81% vs 66% p=0.27, respectively). Viability of cryopreserved hepatocytes after thawing averaged 50%. However thawed hepatocytes had a normal diazepam metabolism (fig 1) and human albumin synthesis was comparable to that of fresh cells (fig 2).FigureFigureCryopreserved human hepatocytes (CHH) were utilized as the biological component of a BAL device for temporaneous support of two patients affected by FHF awaiting for urgent transplant. A total of 3 BAL applications were performed. Ten to 13 billion viable CHH have been loaded in each BAL. No adverse events were recorded and vital functions remained table during the procedures. Biochemical liver functions showed bilirubin and ammonia reduction at the end of each treatment with CHH based BAL. One patient was successfully bridged to emergency OLTx performed after a single BAL; in the second case there was a spontaneous recovery of liver function after two BAL treatment and surgical arterialization of the portal vein. Both patients are still alive and well at home. Conclusions: the recovery of donor human livers unwanted for transplantation at harvesting, allowed isolation and cryopreservation of viable and functionally active human hepatocytes which have been banked and successfully used for clinical application of a BAL device.

Isolation of human hepatocytes from livers rejected for liver transplantation on a national basis: Results of a 2-year experience

The offer of liver transplantation to many patients affected by liver failure is limited by organ shortage. Clinical application of human-based liver cell therapies, such as bioartificial liver and hepatocyte transplantation, might support liver transplantation, allowing more patients to be treated and decreasing mortality in the waiting list. The development of a standardized method of hepatocyte isolation is a mainstay for large-scale application of liver cell therapy. The aim of this study is to analyze retrospectively a 2-year experience of human hepatocyte isolation from livers rejected from transplantation at organ harvesting, performed on a national basis in Italy. All the livers judged unsuitable for transplantation were considered for hepatocyte isolation. Macrosteatosis greater than 60% was the most common reason of refusal, followed by nonviral cirrhosis. Fifty-four organs were used. Human hepatocyte isolation resulted in more that 7 million liver cells/g of tissue digested with 73% ± 14% viability. Steatotic organs gave better results in terms of cell yield than cirrhotic livers. Isolated hepatocytes were able to perform specific liver functions, and evidence of factor IX and albumin messenger RNA (mRNA) production was reported when cells were plated in culture. Modifications of the traditional method of hepatocyte isolation, aimed at reducing ischemia-reperfusion damage and improving post-isolation cell conditions, showed improvements in post-isolation viability. In conclusion, we show that it is possible to use the vast majority of livers not suitable for transplantation on a national basis for human hepatocyte isolation, obtaining a large amount of viable functioning human hepatocytes that might be used for cell transplantation and therapy. (Liver Transpl 2003;9:506-512.)