Use of (18)F-FDG PET/CT to locate primary malignancies in patients with hepatic cirrhosis and malignant ascites.

OBJECTIVE: Ascites in patients with hepatic cirrhosis is caused by cirrhosis in most cases. For most malignant ascites, the primary malignancy could be readily identified using conventional imaging methods, e.g., computed tomography (CT) and magnetic resonance imaging (MRI). However, in a small fraction of the patients, the primary malignancy remains occult even with these examinations. In this retrospective study, we assessed the usefulness of (18)F-FDG PET/CT in patients with hepatic cirrhosis and malignant ascites of otherwise unknown origin. METHODS: Twenty-eight patients with malignant ascites of unknown primary sites after CT, MRI and ultrasound during the period of five years between January 2008 and December 2012 had received (18)F-FDG PET/CT. Medical records of these patients were reviewed and analyzed. RESULTS: Elevated (18)F-FDG absorption was found in 23 of 28 cases in the following sites: gastrointestinal tract (n=10, 43.5%), prostate (n=5, 21.7%), peritoneum (n=4, 13.3%), and ovary (n=4, 13.3%). Cancer was confirmed by pathology in 20 cases after open or laparoscopic surgeries. Five patients were found to have benign ascites, among which, 3 were found to be false positive due to tuberculosis. SUV values were significantly higher for tumors than for benign lesions (mean values, 6.95 vs. 2.94; P=0.005). CONCLUSIONS: The (18)F-FDG PET/CT can be as a powerful imaging tool in identifying tissue origin in liver cirrhosis patients suspected of cancers or with cancers of unknown primary sites.

[Screening and cloning of hepatitis C virus non-structural protein 4A interacting protein gene in hepatocytes].

OBJECTIVE: To investigate biological functions of hepatitis C virus (HCV) non-structural protein 4A (NS4A). METHODS: Yeast-two hybrid technique was performed to seek proteins in hepatocytes interacting with HCV NS4A. HCV NS4A bait plasmid was constructed by ligating the NS4A gene with carrier plasmid pGBKT7, then it was transformed into yeast AH109 (alpha type). The transformed yeast cells were amplified and mated with yeast cells Y187 (alpha type) containing liver cDNA library plasmid pACT2 in 2 x YPDA medium. Diploid yeast cells were plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing X-alpha-gal for selection two times. After extracting plasmid from blue colonies, plasmid DNA was transformed into competent E.coli and analyzed by DNA sequencing and bioinformatics methods. RESULTS: Among twenty-two positive colonies there were eleven positive for metallothionein 2A, three for eukaryotic translation elongation factor 1 alpha 1, two for albumin, two for RNA binding motif protein 21, two for myomesin, one for cytochrome C oxidase II, and one for ATPase. CONCLUSIONS: Genes of HCV NS4A interacting proteins in hepatocytes were successfully cloned and the results pave the way for studying the biological functions of NS4A and associated proteins.

[Expression of human fibroleukin gene acute on chronic hepatitis B and its clinical significance].

OBJECTIVE: To investigate the mRNA and protein expressions of human fibroleukin gene (hfg12) in acute on chronic (AOC) hepatitis B and its clinical significance. METHODS: Liver tissues were obtained from 23 patients with AOC hepatitis B, 13 patients with chronic hepatitis, and 14 patients with cirrhosis to be examined histologically. Immunohistochemistry and in situ hybridization were used to detect the mRNA and protein expressions of hfg12 in the liver tissues. Double staining was used to the hfg12 positive samples to examine both the hfg12 and fibrin. Four specimens of liver tissue from normal donors were used as controls. RESULTS: Immunohistochemistry showed that hfg12 was expressed in the liver tissues of 21 out of the 23 patients with AOC hepatitis B (91.30%) and only one out of the 13 patients with chronic hepatitis (7.69%). In situ hybridization showed that hfg12 was expressed in the liver tissues of 13 out of he 23 patients with AOC hepatitis B and in none of the 27 patients with chronic hepatitis or cirrhosis. In patients with AOC hepatitis Kupffer's cell, CD68 positive, was numerous and big, mainly distributed in the necrosis areas. It was identified as the same of hfg12-expressing cells. CONCLUSION: High expression of hfg12 is one of the molecular mechanisms of necrosis of liver cells in AOC hepatitis.

[Hfgl2/fibroleukin expression in liver and peripheral blood mononuclear cells (PBMC) and its correlation with disease severity].

OBJECTIVE: Viral hepatitis remains a major public health problem and the most common type of liver disease worldwide. There are an increasing number of patients with chronic hepatitis B who develop acute hepatitis on chronic condition (AOC) and die of acute hepatic failure both as a result of lack of understanding of the pathogenesis of the disease and lack of effective treatment. The hallmark of AOC is the extreme rapidity of the necromicroinflammatory process resulting in widespread or total hepatocellular necrosis in weeks or even days. Our previous studies have shown in an experimental animal model of fulminant viral hepatitis caused by murine hepatitis virus strain 3, the importance of macrophage activation, and expression of a unique gene mfgl 2 which encodes a serine protease capable of directly cleaving prothrombin to thrombin, resulting in widespread fibrin deposition within the liver and hepatocyte necrosis. The undergoing study in this report is designed to identify the role of hfgl 2 (human fibrinogen like protein 2) /fibroleukin in patients with viral hepatitis. METHODS: Liver tissues were obtained from 23 patients with AOC hepatitis B, and from 13 patients with inactive chronic hepatitis B (CHB) and 14 patients with chronic hepatitis B with cirrhosis during the year of 1995 to the end of 2001. Liver biopsies were performed within 30 min after the patients were diagnosed with death as a result of acute hepatic failure. Liver samples were also obtained from 4 liver donors as normal controls. In addition, peripheral blood mononuclear cells (PBMC) were isolated from 30 patients (unpaired) with AOC hepatitis B and 10 patients with CHB during the May of 2001 to March of 2002 and 10 healthy volunteer as negative control. PBMCs were freshly isolated and smeared on slides and kept at -80 degree C for further use. Histological sections were stained with hemotoxylin and eosin. A 169 bp of hfgl 2 cDNA probe and a polyclonal or monoclonal antibody against hfgl 2 were used to detect the expression of hfgl 2 mRNA and protein in liver samples as well as PBMC by immune histochemistry separately. RESULTS: Liver tissues from the patients with acute on chronic hepatitis had classical pathological features of acute necroinflammation. Hfgl 2 was detected by immune histochemistry in 21 of 23 patients (91.3%) in liver sections from patients with acute on CHB, while only 1 of 13 patients (7.7%) with CHB and cirrhosis and no evidence of active disease had hfgl 2 mRNA or protein expression. 28 of 30 patients (93.3%) with acute on CHB and 1 of 10 with CHB were detected with hfgl 2 expression in PBMC. There was no hfgl 2 expression in either the liver tissue or the PBMC from the normal donors. There was positive correlation of hfgl 2 expression and the severity of the disease displayed by the value of bilirubin and PT. CONCLUSION: The molecular and cellular results reported here in patients with acute on chronic hepatitis and who died of acute hepatic failure correlates with previous report in 8 patients with fulminant hepatic failure (FHF) and mimic closely the changes observed in the murine model of fulminant viral hepatitis in which the pathogenesis of the disease has been studied in a stepwise fashion. This study further suggests that virus induced hfgl 2 prothrombinase/fibroleukin expression and the potent function of the protein it encodes plays a pivotal role in initiating acute severe hepatitis on the baseline of chronic hepatitis. The measurement of hfgl 2/fibroleukin expression in PBMC may serve as a useful marker to monitor the severity of disease in patients with the AOC hepatitis B and a target for therapeutic intervention.

Sepsis-associated cholestasis in adult patients: a prospective study.

BACKGROUND: Sepsis-associated cholestasis is a common problem in neonatal patients. However, there are limited data related to sepsis-associated cholestasis in adults. In this study, the authors assessed the clinical characteristics, risk factors and outcome of adult patients with sepsis-associated cholestasis. METHODS: An observational prospective single-center study was conducted. A total of 608 patients with sepsis (66 patients with cholestasis and 542 without evidence of cholestasis) from January 1, 2005, to December 31, 2011, were included from the infectious disease unit. Demographic, clinical and laboratory information were recorded on admission for all patients. Additional data were also collected on the day of the 1st episode of bacteremia for patients who developed cholestasis. Accordingly, the organ dysfunction scores (Acute Physiology and Chronic Health Evaluation [APACHE] II and Sequential Organ Failure Assessment [SOFA]) were assessed on the same day. RESULTS: The mean age of the 608 patients was 49.3 ± 11.4 years (range, 22-83 years); 312 (51.3%) patients were men, 296 (48.7%) were women. The mean APACHE II and SOFA score were 15.2 ± 6 and 5.6 ± 2.3, respectively. Sepsis-associated cholestasis was strongly associated with older age, biomarkers of organ dysfunction and clinical composite scores (APACHE II and SOFA). Mortality was higher in patients with sepsis-associated cholestasis (10.6%) compared with subjects with sepsis without cholestasis (1.5%) (P < 0.05). CONCLUSIONS: The authors found that sepsis-associated cholestasis affects the outcome of patients with sepsis in the infectious disease unit. Additional clinical studies are necessary to elucidate the pathology and pathophysiology of sepsis-associated cholestasis.

Hepatitis B Virus Genotype B and High Expression of Interferon Alpha Receptor ß Subunit are Associated With Better Response to Pegylated Interferon Alpha 2a in Chinese Patients With Chronic Hepatitis B Infection.

BACKGROUND: Hepatitis B virus (HBV) is one of leading causes of various hepatic diseases including acute and chronic hepatitis, cirrhosis, and hepatocellular carcinoma. Hundreds of million people worldwide are infected by HBV, chronically. OBJECTIVES: This study in conducted to investigate the influence of Hepatitis B virus (HBV) genotypes and type I IFN-αreceptor ß subunit (IFNAR2) expression in liver on response to treatment with pegylated IFN-α-2a (Peg-IFN-α-2a) for chronic hepatitis B infection. PATIENTS AND METHODS: In this study, 65 eligible patients with chronic hepatitis B disease were enrolled. HBV genotypes of these patients were analyzed by using PCR-RFLP of the surface gene of HBV. The expression of IFNAR2 in the liver was immune histochemically investigated using anti-IFNAR2 antibody. All immune histochemical slides were read semi-quantitatively by image analysis. Chronic hepatitis B patients were treated with Peg-IFN-α2a therapy for a 48-week period and followed up for 24 weeks. Baseline characteristics and sustained viral response (SVR) to Peg-IFN-α-2a therapy were evaluated. RESULTS: 55 % of patients exhibited HBV genotype B and 31.7 % patients exhibited HBV genotypes C infections. After treatment with Peg-IFN-α-2a, SVR was achieved in 66.7 % of patients with HBV genotype B and in 26.3 % of patients with HBV genotype C (P = 0.009). Semiquantitative and the image analysis indicated by gray level values revealed a higher IFNAR2 expression in the group with severe inflammation (P < 0.001). Patients' high IFNAR2 protein expression had a significant impact on SVR to Peg-IFN-α-2a therapy (P = 0.028). CONCLUSIONS: HBV genotype B and high expression of IFNAR2 in the liver of chronic hepatitis B patients are closely associated with better response to Peg-IFN-α-2a therapy in chronic hepatitis B disease.

B-cell clonality in the liver of hepatitis C virus-infected patients.

AIM: The association of hepatitis C virus (HCV) infection with type II mixed cryoglobulinemia is well established, but the role of HCV in B-cell lymphoma remains controversial. In patients with HCV infection, B-cell clonal expansions have been detected in peripheral blood and bone marrow, and a high prevalence of B-cell non-Hodgkin's lymphomas has been documented. Liver biopsies in chronic HCV infection frequently show portal lymphoid infiltrates with features of B follicles, whose clonality has not yet been investigated. The object of this study was to determine the frequency of liver-infiltrating monoclonal B-cells in 40 patients with HCV infection. METHODS: Eight hundred and forty-eight patients were studied prospectively, including 40 HCV-positive patients and 808 patients with chronic hepatitis B virus (HBV) infection. Immunohistochemical study for B- and T-cell markers was performed on the paraffin-embedded liver tissue sections. The clonality of lymphoid B-cells was tested using a polymerase chain reaction (PCR) approach designed to identify immunoglobulin heavy chain gene (IgH) rearrangements. RESULTS: Liver-infiltrating monoclonal B-cells were detected in the liver for 4 (10%) of 40 HCV-positive patients but were present in only 3 (0.37%) of 808 liver biopsy specimens with chronic HBV infection. Chi-square testing showed that the monoclonal B-cells infiltration in the liver was more frequent in the HCV-infected patients (P = 0.000). A clonal IgH rearrangement was detected in 5 (71.4%) of 7 liver biopsy specimens with monoclonal B-cells infiltration. In 2 of 5 patients with both a clonal B-cell expansion and monoclonal B-cells infiltration in the liver, a definite B-cell malignancy was finally diagnosed. CONCLUSION: Liver-infiltrating monoclonal B-cells are detected in the liver of patients with chronic HCV and HBV infection. A high percentage of patients with monoclonal B-cells infiltration and B-cell clonality in the liver were finally diagnosed as having a definite B-cell malignancy.

[Screening and cloning of hepatitis C virus non-structural protein 4B interacting protein gene in hepatocytes].

OBJECTIVE: To investigate biological functions of non-structural protein 4B (NS4B) of hepatitis C virus (HCV), yeast-two hybrid technique was performed to seek proteins in hepatocytes interacting with HCV NS4B. METHODS: HCV NS4B bait plasmid was constructed by ligating the NS4B gene with carrier plasmid pGBKT7 and transformed into yeast cells AH109 (type alpha). The transformed yeast cells were amplified and mated with yeast cells Y187 (alpha type) containing liver cDNA library plasmid pACT2 in 2 x YPDA medium. Diploid yeast cells were plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing x-alpha-gal for selecting two times. After extracting plasmid from blue colonies, plasmid DNA was transformed into competent Escherichia coli and analysed by DNA sequencing and bioinformatics. RESULTS: Five genes in eight positive colonies were obtained. There were one NADH dehydrogenase subunit 3, one cytochrome c oxidase subunit III, one retinol binding protein 4, one reticulon 3-A (RTN3) and one fibrinogen gamma polypeptide (FGG). CONCLUSION: Genes of HCV NS4B interacting proteins in hepatocytes were successfully cloned and the results paved the way for studying the biological functions of NS4B and associated proteins.

Hepatitis C virus may infect extrahepatic tissues in patients with hepatitis C.

AIM:To explore the status of extrahepatic hepatitis C virus (HCV) infection and replication in hepatitis C patients,and its potential implication in HCV infection and pathogenicity.METHODS:By reverse-transcriptase poly-merase chain reaction (RT-PCR),in situ hybridization (ISH) and immunohis-tochemistry, HCV RNA, HCV replicative intermediate (minus-strand of HCV RNA), and HCV antigens were detected in 38 autopsy extrahepatic tissue specimens (including 9 kidneys, 9 hearts, 9 pancreas, 5 intestines, 2 adrenal glands, 2 spleens, 1 lymph node, and 1 gallbladder) from 9 hepatitis C patients, respectively; and the status of HCV replication in extrahepatic tissues was studied.RESULTS:By RT-PCR, all 9 patients were positive for HCV RNA in kidney, heart, pancreas, and intestine, but only 6(66.7%) patients were positive for HCV replicative intermediate. HCV RNA and HCV antigens were detected in kidney, heart, pancreas, intestine, adrenal gland, lymph node, and gallbladder in 5(55.6%) and 6(66.7%) patients by ISH and immuno-histochemistry, respectively. HCV RNA and HCV antigens were not detected in these extrahepatic organs in 3(33.3%) patients, although their livers were positive for HCV.HCV replicative intermediate detected by RT-PCR was consistent with HCV RNA and HCV antigens detected by ISH and immunohistochemistry (Kappa =0.42-0.75). HCV RNA and HCV antigens were detected in myocardial cells, epithelial cells of intestinal gladular, interstitial cells of kidney, epithelial cells of tubules and glomerulus, pancreas acinar cells and epithelial cells of pancreatic duct, epithelial cells of mucous membrane sinus of gallbladder, cortex and medulla cells in adrenal gland,and mononuclear cells in lymph node. HCV RNA was also detected in bile duct epithelial cells, sinusoidal cells, and mononuclear cells in liver tissues by ISH.CONCLUSION:HCV can infect extrahepatic tissues, and many various tissue cells may support HCV replication; extrahepatic HCV infection and replication may be of concomitant state in most of patients with hepatitis C. The infected extrahepatic tissues might act as a reservoir for HCV, and play a role in both HCV persistence and reactivation of infection. HCV as an etiologic agent replicating and expressing viral proteins in extrahepatic tissues itself contributes to extrahepatic syndrome associated-HCV infection in a few patients with chronic HCV infection.