Clonal expansion of hepatocytes with a selective advantage occurs during all stages of chronic hepatitis B virus infection.

Hepatocyte clone size was measured in liver samples of 21 patients in various stages of chronic hepatitis B virus (HBV) infection and from 21 to 76 years of age. Hepatocyte clones containing unique virus-cell DNA junctions formed by the integration of HBV DNA were detected using inverse nested PCR. The maximum hepatocyte clone size tended to increase with age, although there was considerable patient-to-patient variation in each age group. There was an upward trend in maximum clone size with increasing fibrosis, inflammatory activity and with seroconversion from HBV e-antigen (HBeAg)-positive to HBeAg-negative, but these differences did not reach statistical significance. Maximum hepatocyte clone size did not differ between patients with and without a coexisting hepatocellular carcinoma. Thus, large hepatocyte clones containing integrated HBV DNA were detected during all stages of chronic HBV infection. Using laser microdissection, no significant difference in clone size was observed between foci of HBV surface antigen (HBsAg)-positive and HBsAg-negative hepatocytes, suggesting that expression of HBsAg is not a significant factor in clonal expansion. Laser microdissection also revealed that hepatocytes with normal-appearing histology make up a major fraction of the cells undergoing clonal expansion. Thus, preneoplasia does not appear to be a factor in the clonal expansion detected in our assays. Computer simulations suggest that the large hepatocyte clones are not produced by random hepatocyte turnover but have an as-yet-unknown selective advantage that drives increased clonal expansion in the HBV-infected liver.

Awareness and opinions of non-alcoholic fatty liver disease by hospital specialists.

BACKGROUND/AIM: Subjects with metabolic risk factors for non-alcoholic fatty liver disease (NAFLD) are commonly seen by hospital specialists other than gastroenterologists/hepatologists. The aim of this study was to assess the awareness of NAFLD and opinions regarding management among non-hepatologists at two major tertiary hospitals in Brisbane. METHODS: A face-to-face questionnaire assessing current beliefs and practices regarding NAFLD was administered to specialists and specialists-in-training across six specialties (internal medicine, cardiology/cardiac surgery, endocrinology, thoracic medicine, rheumatology and nephrology). RESULTS: One hundred clinicians were surveyed with 99% returning completed questionnaires (>89% questions answered). The majority of respondents (75%) believe the prevalence of NAFLD in the general population to be ≤ 10%, although two-thirds feel that its incidence will rise markedly. The vast majority (>90%) appreciate that traditional cardiovascular risk factors (obesity, hypertriglyceridaemia and diabetes) are risk factors for NAFLD and acknowledge that these are common in non-hepatology patients. Despite this, most believe that NAFLD is uncommon in their own patients (89% indicated a prevalence ≤ 30%). The vast majority (93%) agree that non-alcoholic steatohepatitis (NASH) is associated with increased overall mortality, but 60% also believe that simple steatosis confers increased liver-related mortality. Most (74%) agree that a diagnosis of NASH cannot be made using liver enzymes, but 67% support 6-monthly liver function tests as the most effective way to monitor progression of NAFLD. Most respondents (71%) make no referrals to hepatology for suspected NAFLD. CONCLUSIONS: Non-hepatologists appreciate the seriousness of NAFLD but appear to underestimate its prevalence, especially among their own patients despite known risk factors. Attitudes regarding simple steatosis versus NASH and appropriate monitoring of suspected NAFLD suggest that more can be done to improve the understanding of this disease among non-hepatologists. This has implications for targeting 'at-risk' populations and appropriate referral of patients to hepatology clinics.

Gastric adenocarcinoma and proximal polyposis of the stomach (GAPPS): a new autosomal dominant syndrome.

OBJECTIVE: The purpose of this study was the clinical and pathological characterisation of a new autosomal dominant gastric polyposis syndrome, gastric adenocarcinoma and proximal polyposis of the stomach (GAPPS). METHODS: Case series were examined, documenting GAPPS in three families from Australia, the USA and Canada. The affected families were identified through referral to centralised clinical genetics centres. RESULTS: The report identifies the clinical and pathological features of this syndrome, including the predominant dysplastic fundic gland polyp histology, the exclusive involvement of the gastric body and fundus, the apparent inverse association with current Helicobacter pylori infection and the autosomal dominant mode of inheritance. CONCLUSIONS: GAPPS is a unique gastric polyposis syndrome with a significant risk of gastric adenocarcinoma. It is characterised by the autosomal dominant transmission of fundic gland polyposis, including areas of dysplasia or intestinal-type gastric adenocarcinoma, restricted to the proximal stomach, and with no evidence of colorectal or duodenal polyposis or other heritable gastrointestinal cancer syndromes.

Successful immunotherapy of HCMV disease using virus-specific T cells expanded from an allogeneic stem cell transplant recipient.

Opportunistic infection remains the principal cause of mortality in allogeneic stem cell transplant recipients with active extensive chronic graft-versus-host disease. Human cytomegalovirus (HCMV) represents an important cause of disease in this setting and the toxicity of protracted and recurrent antiviral treatment together with eventual drug resistance represents a significant limitation to therapy. Although the expansion and adoptive transfer of HCMV-specific T cells from the healthy original donor can be an effective strategy to control viral replication, this is not possible when donors are seronegative or are subsequently inaccessible. Here we demonstrate for the first time, the successful expansion of HCMV-specific T cells from a seropositive transplant recipient of a seronegative graft with active HCMV disease and the long-term reconstitution of protective antiviral immunity following their adoptive transfer back into the patient.

Liver fluke-associated and sporadic cholangiocarcinoma: an immunohistochemical study of bile duct, peribiliary gland and tumour cell phenotypes.

AIM: To compare cell phenotypes displayed by cholangiocarcinomas and adjacent bile duct lesions in patients from an area endemic in liver-fluke infestation and those with sporadic cholangiocarcinoma. METHODS: 65 fluke-associated and 47 sporadic cholangiocarcinomas and 6 normal livers were studied. Serial paraffin-wax sections were stained immunohistochemically with monoclonal antibodies characterising a Brunner or pyloric gland metaplasia cell phenotype (antigens D10 and 1F6), intestinal goblet cells (antigen 17NM), gastric foveolar apomucin (MUC5AC), a gastrointestinal epithelium cytokeratin (CK20) and the p53 protein. RESULTS: 60% of the 112 cholangiocarcinomas expressed antigen D10, 68% MUC5AC, 33% antigen 17NM and 20% CK20; 37% showed overexpression of p53. When present together in a cholangiocarcinoma, cancer cells expressing D10 were distinct from those displaying 17NM or MUC5AC. Many more fluke-associated cholangiocarcinomas than sporadic cholangiocarcinomas displayed 17NM and p53 expression. Most cases of hyperplastic and dysplastic biliary epithelium expressed D10 strongly. Pyloric gland metaplasia and peribiliary glands displayed D10 and 1F6, with peribiliary gland hyperplasia more evident in the livers with fluke-associated cholangiocarcinoma; goblet cells in intestinal metaplasia stained for 17NM. No notable association of expression between any two antigens (including p53) was found in the cancers. CONCLUSIONS: Most cases of dysplastic biliary epithelium and cholangiocarcinoma display a Brunner or pyloric gland cell phenotype and a gastric foveolar cell phenotype. The expression of D10 in hyperplastic and dysplastic epithelium and in cholangiocarcinoma is consistent with a dysplasia-carcinoma sequence. Many more fluke-associated cholangiocarcinomas than sporadic cholangiocarcinoma display an intestinal goblet cell phenotype and overexpress p53, indicating differences in the aetiopathology of the cancers in the two groups of patients.

A combination of genetic polymorphisms increases the risk of progressive disease in chronic hepatitis C.

BACKGROUND: There is increasing interest in the influence of host genetic factors on hepatic fibrosis, and whether genetic markers can reliably identify subjects at risk of developing severe disease. We hypothesised that hepatitis C virus (HCV) infected subjects with progressive fibrosis, classified using strict criteria based on histology at biopsy in addition to disease duration would be more likely to inherit several genetic polymorphisms associated with disease progression compared with subjects with a low rate of disease progression. METHODS: We examined polymorphisms in eight genes that have been reported to have an association with hepatic fibrosis. RESULTS: Associations between polymorphisms in six genes and more rapidly progressing fibrosis were observed, with individual adjusted odds ratios ranging from 2.1 to 4.5. The relationship between rapidly progressing fibrosis and possession of > or =3, > or =4, or > or =5 progression associated alleles was determined and the adjusted odds ratios increased with increasing number of progression associated alleles (9.1, 15.5, and 24.1, respectively). Using logistic regression analysis, a predictive equation was developed and tested using a second cohort of patients with rapidly progressing fibrosis. The predictive equation correctly classified 80% of patients in this second cohort. CONCLUSIONS: This approach may allow determination of a genetic profile predictive of rapid disease progression in HCV and identify patients warranting more aggressive therapeutic management.

Eosinophilic enteritis in northeastern Australia. Pathology, association with Ancylostoma caninum, and implications.

While eosinophilic gastroenteritis is considered a rare condition, eosinophilic enteritis without gastric involvement is quite common in northeastern Australia. We present 79 patients with biopsy-proven eosinophilic enteritis, 70 seen since 1987. In 10 patients, eosinophilic enteritis was associated with infection by single, sexually immature, adult hookworms, most positively identified as the common dog hookworm Ancylostoma caninum. An additional 22 patients (of 34 tested) had serological evidence of A. caninum exposure. The essential pathology, i.e., edema and eosinophilic infiltration of the gut wall, ascites, and regional lymphadenopathy, was identical to that seen in eosinophilic gastroenteritis. Additional, more specific features included pathological reactions centered on attached worms, mucosal alterations and ulcers considered to be hookworm bite sites, and submucosal and lymph node granulomas with central eosinophil degranulation and degradation products. Since A. caninum has an almost worldwide distribution, it is probable that A. caninum-induced eosinophilic enteritis occurs outside Australia. We show that the worm is easily overlooked in pathological specimens and that care is required to preserve worms intact for specific parasitological identification. The clinical and pathological features were similar to those seen in another human enteric helminthic zoonosis, anisakiasis. The possibility that there are yet other undiscovered intestinal zoonoses remains.

Persistence of donor-reactive CD4+ T cells in liver and spleen of rats tolerant to a liver allograft.

BACKGROUND: In the rat, orthotopic liver transplantation from a DA strain donor to a PVG recipient causes an early rejection response that spontaneously resolves over the following weeks to yield long-lasting, donor-specific tolerance. METHODS: Limiting dilution analysis was used to estimate the frequencies of host CD4+ cells able to proliferate in response to donor antigens in the grafted liver and spleen of recipients during and after tolerance induction. RESULTS: Compared with naive PVG rats, both the frequencies and absolute numbers of donor-reactive host CD4+ cells in the liver and spleen rose significantly during the first week after transplantation and remained elevated for at least 3 months. CONCLUSION: We conclude that the development of tolerance in this model is not associated with deletion of clonogenic donor-reactive CD4+ T cells by clonal exhaustion or any other mechanism.

Clinicopathological analysis of liver allograft biopsies with late centrilobular necrosis: a comparative study in 54 patients.

BACKGROUND: Centrilobular necrosis (CLN) in liver allografts can be a difficult lesion to interpret histologically. Although long recognized in association with developing chronic rejection, recent studies have described the lesion in association with a number of other disease processes. To clarify the histologic features that could allow a specific diagnosis to be made and to determine the outcome in different diagnostic groups, we assessed biopsies from 54 patients with CLN. METHODS: Biopsies were classified as CLN with acute cellular rejection (ACR), CLN with hepatitis, CLN with developing chronic rejection (CR), and CLN of other etiology. Histologic features were assessed and then compared between groups, and clinical outcomes were noted. RESULTS: Discriminating features for the different groups were as follows: CLN and ACR showed bile duct injury, endothelialitis, and acinar congestion. CLN and CR showed severe bile duct injury, bile duct loss, or centrilobular swelling. CLN and hepatitis was often a diagnosis of exclusion, although interface hepatitis was more common in this group. Cases of autoimmune hepatitis usually demonstrated plasma cell predominance in the portal and acinar inflammatory infiltrate. Significantly, there was considerable overlap in the histologic features between the groups, accounting for the diagnostic difficulty. Patients in whom the CLN was associated with CR or vascular complications generally required retransplantation or died, but in the groups with ACR and hepatitis, the outcome was more favorable. CONCLUSIONS: With regard to most liver allograft biopsies showing late CLN, it is possible to make a specific diagnosis despite overlapping histologic features; this allows specific therapy to be instituted. Ultimately this is likely to contribute to improved graft survival.

Role of donor leukocyte chimerism in establishing the etiology of neutropenia after liver transplantation.

BACKGROUND: The quantitation of donor leukocyte chimerism may aid in establishing the etiology of neutropenia after liver transplantation. METHODS: The incidence and clinical and laboratory characteristics of severe neutropenia were studied in adults who have undergone liver transplantation at our institution over the last 4 years. RESULTS: Severe neutropenia developed in 5 of 156 patients (3%). The clinical and pathological features were nonspecific. In two patients with a delayed diagnosis of graft-versus-host disease (GVHD), donor leukocytes comprised > or = 50% of the circulating peripheral blood mononuclear cells. In a third patient, an earlier diagnosis of GVHD was suspected on the basis of a donor leukocyte count of 3-10% in the peripheral blood. In contrast, donor leukocyte chimerism was < or = 0.01% in two patients with probable drug-induced neutropenia CONCLUSIONS: The determination of donor leukocyte chimerism has an important role in the investigation of neutropenia after liver transplantation, allowing early diagnosis and treatment of GVHD.

Timely topic: Premalignant lesions associated with adenocarcinoma of the upper gastrointestinal tract.

The changing incidence of adenocarcinomas, particularly in the oesophagus and gastric cardia, has led to the rapid expansion of screening programmes aimed at detecting the precursor lesion of dysplasia before adenocarcinoma develops. The pathologist now has an important role in first diagnosing patients at risk for developing dysplasia, and then correctly classifying dysplasia when it occurs. Barrett's oesophagus has had different diagnostic criteria in previous years but is currently diagnosed by the presence of intestinal metaplasia of any length in the true oesophagus. Intestinal metaplasia confined only to the gastro-oesophageal junction or cardia is of uncertain significance but is probably common, with less risk of progressing to dysplasia or malignancy. In the stomach, patients with autoimmune atrophic gastritis and Helicobacter-associated multifocal atrophic gastritis have an increased risk of adenocarcinoma, but screening protocols are not well-developed compared with those used for Barrett's oesophagus. Dysplasia of glandular epithelium can be classified using well-described criteria. Low grade dysplasia is the most common type and regresses or remains stable in the majority of patients. High grade dysplasia is more ominous clinically, with a propensity to coexist with or progress to adenocarcinoma.

Fibrolamellar hepatocellular carcinoma: a case report.

A 6-year-old Malay boy presented with fever and abdominal pain for 2 months. Computerised tomography showed a nodular mass in the left lobe of the liver. There was also portal vein thrombosis on the left side. Serum alpha-fetoprotein was not elevated and Hepatitis B antigen was negative. Biopsy of the liver mass led to a histological diagnosis of fibrolamellar hepatocellular carcinoma. In view of extensive tumour involvement, he could not be operated on but was treated with chemotherapy. However, the tumour did not respond. While this is expected for fibrolamellar hepatocellular carcinoma, the possibility of the tumour having a component of ordinary hepatocellular carcinoma could not be excluded as the tumour was not resected. Fibrolamellar hepatocellular carcinoma is a rare histological subtype of hepatocellular carcinoma, associated with a better prognosis. It affects the younger age group and has no association with cirrhosis, hepatitis B virus infection or exposure to oral contraceptives, all of which are implicated in ordinary hepatocellular carcinoma. Serum alpha-fetoprotein level is usually within normal limits and other laboratory values are not contributory to the diagnosis. The diagnosis is usually suggested by radiographic studies viz. CT scan of the abdomen, which would show an irregular non-homogenous mass in the liver, and confirmed by histological examination. The most characteristic microscopical feature is fibrosis arranged in a lamellar fashion around polygonal and deeply eosinophilic neoplastic hepatocytes.

Non-response to antiviral therapy is associated with obesity and increased hepatic expression of suppressor of cytokine signalling 3 (SOCS-3) in patients with chronic hepatitis C, viral genotype 1.

BACKGROUND: Interferon alpha (IFN-alpha) activated cellular signalling is negatively regulated by inhibitory factors, including the suppressor of cytokine signalling (SOCS) family. The effects of host factors such as obesity on hepatic expression of these inhibitory factors in subjects with chronic hepatitis C virus (HCV) are unknown. OBJECTIVES: To assess the independent effects of obesity, insulin resistance, and steatosis on response to IFN-alpha therapy and to determine hepatic expression of factors inhibiting IFN-alpha signalling in obese and non-obese subjects with chronic HCV. METHODS: A total of 145 subjects were analysed to determine host factors associated with non-response to antiviral therapy. Treatment comprised IFN-alpha or peginterferon alpha, either alone or in combination with ribavirin. In a separate cohort of 73 patients, real time-polymerase chain reaction was performed to analyse hepatic mRNA expression. Immunohistochemistry for SOCS-3 was performed on liver biopsy samples from 38 patients with viral genotype 1 who had received antiviral treatment. RESULTS: Non-response (NR) to treatment occurred in 55% of patients with HCV genotypes 1 or 4 and 22% with genotypes 2 or 3. Factors independently associated with NR were viral genotype 1/4 (p < 0.001), cirrhosis on pretreatment biopsy (p = 0.025), and body mass index > or = 30 kg/m2 (p = 0.010). Obese subjects with viral genotype 1 had increased hepatic mRNA expression of phosphoenolpyruvate carboxy kinase (p = 0.01) and SOCS-3 (p = 0.047), in comparison with lean subjects. Following multivariate analysis, SOCS-3 mRNA expression remained independently associated with obesity (p = 0.023). SOCS-3 immunoreactivity was significantly increased in obesity (p = 0.013) and in non-responders compared with responders (p = 0.014). CONCLUSIONS: In patients with chronic HCV viral genotype 1, increased expression of factors that inhibit interferon signalling may be one mechanism by which obesity reduces the biological response to IFN-alpha.

Characterization of tumour-infiltrating lymphocytes and apoptosis in colitis-associated neoplasia: comparison with sporadic colorectal cancer.

The development of colorectal cancer is a major complication for patients with chronic idiopathic colitis. Colitis-associated tumours tend to occur at a younger age and be more aggressive than sporadic colorectal cancers. While we have previously associated the presence of tumour-infiltrating lymphocytes (TILs) and increased apoptosis in sporadic colorectal cancer with high-level microsatellite instability and improved prognosis, little is known of the relationship between these variables in colitis-associated colorectal cancer. The aim of this study was to correlate TILs and tumour cell apoptosis in colitis-associated neoplasms stratified according to microsatellite instability. Twenty tumour and 11 dysplastic samples resected from 21 patients with long-standing colitis were analysed for microsatellite instability at 10 microsatellite markers. TIL distribution (CD3, CD8) and function (granzyme B) were quantified by immunohistochemistry. Neoplastic cell apoptosis was assessed using the M30 CytoDEATH antibody. These findings were compared with 40 microsatellite stable (MSS) sporadic colorectal cancers previously evaluated for TILs and neoplastic apoptosis. Low-level microsatellite instability was found in 1/20 colitis-associated tumours. All other colitis-associated lesions were designated MSS. CD3(+) and CD8(+) TIL counts were significantly higher in colitis-associated lesions compared with MSS sporadic colorectal cancer (p < 0.0001, p = 0.001 respectively). Despite their higher TIL density, colitis-associated tumours were more likely to present late (Dukes' stage C or D) (p = 0.02). Functionally, colitis-associated TILs demonstrated significantly less granzyme B expression compared to sporadic cancers (p = 0.002). The level of tumour cell apoptosis was similar between the two groups (sporadic, 1.53%; colitis cancers, 1.45%). In conclusion, MSS colitis-associated tumours have a higher prevalence of CD3(+)/CD8(+) TILs but no associated increase in tumour cell killing by apoptosis. Unlike cytotoxic T cells in sporadic colorectal cancer, TILs do not appear to enhance the prognosis of colitis-associated colorectal cancer. This may be related to an impairment of granzyme B expression within these lesions.

Nonalcoholic fatty liver disease: is all the fat bad?

Nonalcoholic fatty liver disease is now a major cause of liver disease in developed countries, largely as a result of an epidemic of obesity, diabetes and sedentary lifestyles. This has resulted in raised clinical awareness and diagnostic refinement. The entity encompasses several histologic patterns from benign steatosis to nonalcoholic steatohepatitis, the latter having a significant risk of progressive fibrosis and the development of cirrhosis. Laboratory tests and imaging are not able to distinguish steatosis from steatohepatitis, which requires liver biopsy. However following an assessment of several risk factors, patients can be stratified for the potential risk of fibrosis, allowing the rational use of liver biopsy. This review will describe the various patterns of nonalcoholic fatty liver disease and relate this to disease pathogenesis and progression. Strategies for management, including experimental interventions, will be discussed.

Nephrogenic metaplasia: long-term haemodialysis and anuria as potential risk factors and reversibility with renal transplantation.

AIM: To determine the incidence of nephrogenic metaplasia in patients with defunctionalized bladders due to long-term end-stage renal failure on haemodialysis. METHODS: From the dialysis registry of the Princess Alexandra Hospital 13 anuric patients who had been on haemodialysis for 10 or more years were identified. Of these, seven were currently awaiting renal transplantation and six had successful transplants. Endoscopic assessment of their lower urinary tracts was performed. RESULTS: Three cases of nephrogenic metaplasia were identified. In one case this led to significant haematuria following transplantation. However, the changes of nephrogenic metaplasia disappeared over the subsequent 18 months without specific treatment. CONCLUSIONS: It appears that a chronically defunctionalized bladder in the presence of end-stage renal failure is associated with an increased risk of nephrogenic metaplasia. These changes may spontaneously regress following transplantation and restoration of bladder function.