An OX40/OX40L interaction directs successful immunity to hepatitis B virus.

Depending on age of acquisition, hepatitis B virus (HBV) can induce a cell-mediated immune response that results in either cure or progressive liver injury. In adult-acquired infection, HBV antigens are usually cleared, whereas in infancy-acquired infection, they persist. Individuals infected during infancy therefore represent the majority of patients chronically infected with HBV (CHB). A therapy that can promote viral antigen clearance in most CHB patients has not been developed and would represent a major health care advance and cost mitigator. Using an age-dependent mouse model of HBV clearance and persistence in conjunction with human blood and liver tissue, we studied mechanisms of viral clearance to identify new therapeutic targets. We demonstrate that age-dependent expression of the costimulatory molecule OX40 ligand (OX40L) by hepatic innate immune cells is pivotal in determining HBV immunity, and that treatment with OX40 agonists leads to improved HBV antigen clearance in young mice, as well as increased strength of T cell responses in young mice and adult mice that were exposed to HBV when they were young and developed a CHB serological profile. Similarly, in humans, we show that hepatic OX40L transcript expression is age-dependent and that increased OX40 expression on peripheral CD4+ T cells in adults is associated with HBV clearance. These findings provide new mechanistic understanding of the immune pathways and cells necessary for HBV immunity and identify potential therapeutic targets for resolving CHB.

Age-dependent hepatic lymphoid organization directs successful immunity to hepatitis B.

Hepatitis B virus (HBV) is a major human pathogen that causes immune-mediated hepatitis. Successful immunity to HBV is age dependent: viral clearance occurs in most adults, whereas neonates and young children usually develop chronic infection. Using a mouse model of HBV infection, we sought mechanisms underpinning the age-dependent outcome of HBV and demonstrated that hepatic macrophages facilitate lymphoid organization and immune priming within the adult liver and promote successful immunity. In contrast, lymphoid organization and immune priming was greatly diminished in the livers of young mice, and of macrophage-depleted adult mice, leading to abrogated HBV immunity. Furthermore, we found that CXCL13, which is involved in B lymphocyte trafficking and lymphoid architecture and development, is expressed in an age-dependent manner in both adult mouse and human hepatic macrophages and plays an integral role in facilitating an effective immune response against HBV. Taken together, these results identify some of the immunological mechanisms necessary for effective control of HBV.

KIR genotypic diversity can track ancestries in heterogeneous populations: a potential confounder for disease association studies.

Killer cell immunoglobulin-like receptors (KIR) are encoded by highly polymorphic genes that regulate the activation of natural killer (NK) cells and other lymphocyte subsets and likely play key roles in innate and adaptive immunity. Association studies increasingly implicate KIR in disease predisposition and outcome but could be confounded by unknown KIR genetic structure in heterogeneous populations. To examine this, we characterized the diversity of 16 KIR genes in 712 Northern Californians (NC) stratified by self-assigned ethnicities and compared the profiles of KIR polymorphism with other US and global populations using a reference database. Sixty-eight distinct KIR genotypes were characterized: 58 in 457 Caucasians (NCC), 17 in 47 African Americans (NCAA), 21 in 80 Asians (NCA), 20 in 74 Hispanics (NCH), and 18 in 54 "other" ethnicities (NCO). KIR genotype patterns and frequencies in the 4 defined ethnicities were compared with each other and with 34 global populations by phylogenetic analysis. Although there were no population-specific genotypes, the KIR genotype frequency patterns faithfully traced the ancestry of NCC, NCAA, and NCA but not of NCH whose ancestries are known to be more heterogeneous. KIR genotype frequencies can therefore track ethnic ancestries in modern urban populations. Our data emphasize the importance of selecting ethnically matched controls in KIR-based studies to avert spurious associations.

Peginterferon alfa-2b and weight-based or flat-dose ribavirin in chronic hepatitis C patients: a randomized trial.

UNLABELLED: This prospective, multicenter, community-based and academic-based, open-label, investigator-initiated, U.S. study evaluated efficacy and safety of pegylated interferon (PEG-IFN) alfa-2b plus a flat or weight-based dose of ribavirin (RBV) in adults with chronic hepatitis C. Patients (n = 5027) were randomly assigned to receive PEG-IFN alfa-2b 1.5 microg/kg/week plus flat-dose (800 mg/day) or weight-based (800-1400 mg/day) RBV for 48 weeks (patients with genotype 1, 4, 5, or 6) and for 24 or 48 weeks (genotype 2/3 patients). Primary end point was sustained virologic response (undetectable [<125 IU/mL] serum hepatitis C virus RNA at 24-week follow-up). Sustained virologic response, but not end-of-treatment, rates were significantly higher with weight-based than with flat-dose RBV (44.2% versus 40.5%; P = 0.008). Sustained virologic response rates by intention-to-treat analysis were 34.0% and 28.9%, respectively, in genotype 1 patients (P = 0.005) and 31.2% and 26.7%, respectively, in genotype 1 patients with high baseline viral load (P = 0.056). In genotype 2/3 patients, rates were not significantly different (61.8% and 59.5%, respectively) regardless of treatment duration. Besides greater hemoglobin reductions with weight-based RBV, safety profiles were similar across RBV dosing groups, including the 1400-mg/day group. CONCLUSION: PEG-IFN alfa-2b plus weight-based RBV is more effective than flat-dose RBV, particularly in genotype 1 patients, providing equivalent efficacy across all weight groups. RBV 1400 mg/day is appropriate for patients 105 to 125 kg. For genotype 2/3 patients, 24 weeks of treatment with flat-dose RBV is adequate; no evidence of additional benefit of extending treatment to 48 weeks was demonstrated.

Efficacy of interferon alpha-2b induction therapy before retreatment for chronic hepatitis C.

BACKGROUND/AIMS: Chronic hepatitis C (HCV) patients who have failed previous treatment have low sustained viral response (SVR) rates with repeat treatment. We evaluated whether interferon (IFN) induction during retreatment improves response rates. METHODS: Two randomized, controlled trials were conducted in chronic HCV patients who failed IFN. In Study 1, patients received IFN 3 MU daily plus ribavirin (RBV) 1000 mg/day for 4 weeks, followed by IFN 3 MU TIW plus RBV 1000 mg/day for 44 weeks (induction; n=232), or IFN 3 MU TIW plus RBV 1000 mg/day for 48 weeks (non-induction; n=237). In Study 2, patients received IFN 5 MU B.I.D. plus RBV 1000-1200 mg/day for 2 weeks, followed by pegylated IFN (PEG-IFN) 75-150 mug weekly plus RBV 1000-1200 mg/day for 46 weeks (induction; n=201), or PEG-IFN 75-150 mug weekly plus RBV 1000-1200 mg/day for 48 weeks (non-induction; n=206). The primary end point for both trials was SVR. RESULTS: Induction did not increase SVR compared with non-induction, but did increase the on-treatment response among genotype non-1 patients in Study 2. By intention-to-treat (ITT) analysis, SVR in Study 1 was 13% for induction vs. 9% for non-induction (P=NS). In Study 2 (ITT), SVR was 20% for induction vs. 24% for non-induction (P=NS). However, by non-ITT analysis of Study 2, genotype non-1-previous non-responders showed significantly higher response rates with induction than non-induction. CONCLUSION: For chronic HCV patients who have failed IFN, induction with retreatment does not improve SVR, but may be beneficial for patients with genotype non-1 HCV.

Mycophenolate mofetil in autoimmune hepatitis patients not responsive or intolerant to standard immunosuppressive therapy.

BACKGROUND & AIMS: The immunosuppressive treatment for autoimmune hepatitis (AIH) patients is prednisone and azathioprine. Ten percent to 20% of patients do not respond or are intolerant of standard treatment. The aim of this study was to assess the biochemical, histologic, and hematologic parameters during mycophenolate mofetil (MMF) treatment in AIH patients who did not respond to or were intolerant of prednisone and/or azathioprine. METHODS: A retrospective study was performed of 15 AIH patients who received MMF either as monotherapy or in combination with prednisone after failure or intolerance of the initial regimen. Records were reviewed as to initial therapy, reasons why MMF was initiated, liver enzyme levels, histology on MMF, and complications. RESULTS: The mean age was 60 +/- 15 years. All patients were started on MMF at 1 gram twice a day, 3 on MMF monotherapy, and 12 on prednisone and MMF. The average MMF treatment duration was 41 months. Alanine aminotransferase levels decreased significantly from 91.73 +/- 88.69 to 60.87 +/- 71.2 (P = .03) on MMF treatment. Inflammatory scores (2.59 +/- 0.97 to 1.14 +/- 1.21, P = .02) and Ishak fibrosis scores (4.10 +/- 1.37 to 2.5 +/- 1.51, P = .02) also decreased. No significant hematologic complications were noted during MMF treatment. CONCLUSIONS: Administration of MMF, either as monotherapy or in combination with prednisone, results in biochemical and histologic improvement in AIH patients who are prednisone and/or azathioprine intolerant or resistant without the development of significant complications. MMF should be studied prospectively as an alternative agent in the treatment of autoimmune liver disease.

A two-step process for cytokine production revealed by IL-4 dual-reporter mice.

To monitor IL-4 expression at the single-cell level, we generated mice with insertions of different reporter genes into both copies of the Il4 gene that permitted the simultaneous analysis of IL-4 transcripts via GFP and IL-4 protein secretion by use of huCD2. Innate and adaptive cells competent for IL-4 production were marked by GFP, while cells that presently or recently secreted IL-4 additionally displayed huCD2. After challenge with the strictly enteric helminth, Heligmosomoides polygyrus, GFP-positive innate and adaptive cells disseminated widely, but IL-4 secretion was predominantly mediated by CD4+ T cells in the intestines and draining lymphoid organs. IL-4-competent cells persisted in cured animals, and memory responses reflected rapid cytokine production at the site of rechallenge. These data reveal a two-step process for cytokine production: the first generating poised cells that disseminate systemically and the second inducing the rapid production of the cytokine in response to local stimulation.

Quantitation of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) in the liver of HBV-infected patients by LightCycler real-time PCR.

Antivirals for hepatitis B virus (HBV) reduce viral load and improve liver histology, however, their effect on covalently closed circular DNA (cccDNA), the HBV transcriptional template, has not been extensively examined. This study evaluated a newly designed LightCycler based quantitative cccDNA PCR assay. A linear range of 2.5 x 10(1) to 1 x 10(9) copies/assay using primers specific for HBV cccDNA and 2.5 x 10(1) to 2.5 x 10(9) copies/assay using primers specific for total HBV DNA (tDNA) was established. beta-Globin was used to estimate the number of cells in each PCR reaction. Enzymatic digestion with an ATP-dependent DNase improved the analytic specificity to a greater than 1:10000 ratio of cccDNA:RC DNA (relaxed circular DNA). One-tenth of the extracted DNA from 1mg of liver biopsy, was analyzed from six patients, three HBV-infected and three uninfected individuals, under blinded conditions; three were found positive and three negative for cccDNA and tDNA. Approximately 6 x 10(3) copies of cccDNA/mg of tissue were detected in a pre-transplant biopsy from an HBV-infected patient treated with lamivudine. Sequential post-transplant liver biopsies were negative for both HBV cccDNA and tDNA. An HBV-infected patient with cirrhosis who was antiviral therapy naïve had 3.7 x 10(4) copies of cccDNA/mg of liver tissue. Another treatment-naïve patient with a history of high HBV viral load had 1 x 10(5) copies of cccDNA/mg of tissue (4 x 10 (6) copies of tDNA/mg of tissue). Further studies are warranted but the high level of sensitivity, specificity, rapidity and accuracy provided by this novel assay with the LightCycler system indicate that it could be useful for monitoring antiviral therapy.

De novo non-alcoholic fatty liver disease following orthotopic liver transplantation.

Non-alcoholic fatty liver disease (NAFLD) is an increasingly recognized clinico-pathologic entity typically associated with obesity, type II diabetes and hyperlipidemia. It has been noted to recur after orthotopic liver transplantation (OLT). We report four patients who developed de novo NAFLD within 3 months of OLT without the typical predisposing factors of diabetes mellitus or obesity. Three of the four patients underwent OLT for hepatitis C-related cirrhosis, and the other for alcoholic cirrhosis. Examination of the liver explants revealed no evidence of steatosis. No surreptitious alcohol use or a drug-induced process could be identified in these patients. Treatment of recurrent hepatitis C infection in one patient with interferon and ribavirin led to sustained suppression of the viral RNA to undetectable levels, but no improvement in histology or liver enzymes. All four patients had histologic evidence of preservation injury on the initial post-OLT biopsies, but the significance of this finding in relationship to the development of NAFLD is unknown. NAFLD can develop without any of the known predisposing conditions after transplantation, and this raises further questions about the pathogenesis of this condition.