Volker Meier

Interferon type I gene expression in chronic hepatitis C

Hepatitis C virus (HCV) frequently causes chronic liver disease. The cause of viral persistence might be an inappropriate type I interferon (IFN) induction. To analyze the hosts IFN response in chronic hepatitis C, we measured the transcription level of type I IFN genes as well as type I IFN-regulated genes in liver tissue and corresponding blood samples from patients with chronic hepatitis C, nonviral liver diseases, and a suspected but later excluded liver disease. Competitive and real-time RT-PCR assays were used to quantify the messenger RNA (mRNA) levels of all known IFN-α, IFN-β, and IFN-λ genes and those of some IFN-regulated genes. We failed to detect any hepatic type I IFN mRNA induction, although liver tissue of chronic hepatitis C patients contained high numbers of some type I IFN-inducible effector mRNA molecules. Analysis of peripheral blood samples, however, showed a clear type I IFN induction. Parallel experiments employing HCV replicon cell lines revealed that replication of HCV RNA is not sufficient to induce any type I IFN nor to induce directly type I IFN-regulated genes such as MxA. In conclusion, our data provide evidence for the absence of an induction of type I IFN genes by HCV in the human liver and argue for a further development of type I IFN-based therapies.

Identification of mutations in the ALD-gene of 20 families with adrenoleukodystrophy/adrenomyeloneuropathy.

Adrenoleukodystrophy (ALD), an X-linked inherited metabolic disorder, is the most frequent inborn peroxisomal disease. It leads to demyelination in the central and peripheral nervous system. Defective β-oxidation of saturated very long chain fatty acids (VLCFAs; C22:0–C26:0) in peroxisomes has been shown to lead to an accumulation of VLCFAs in leukoid areas of the central nervous system, peripheral nerves, adrenal gland, and blood. The ALD gene has been recently identified and encodes a 745-amino-acid protein. We screened patients with adrenoleukodystrophy/adrenomyeloneuropathy (ALD/AMN) from 20 kindreds for mutations in the ALD gene. Eleven missense and two nonsense mutations, five deletions, and one insertion were detected by direct sequencing of eight reverse transcribed fragments of the ALD-gene mRNA. Four mutations could be shown to be de novo. All mutations could be confirmed in carriers by sequencing genomic DNA. No correlation between the type of mutation and the severity of the phenotype could be observed. The mutations were not detected in the ALD gene of 30 healthy persons.

MxA gene expression in peripheral blood mononuclear cells from patients infected chronically with hepatitis C virus treated with interferon-α

Hepatitis C virus (HCV) infection causes acute and often chronic liver disease. The treatment of choice is interferon‐α (IFN‐α). The proportion of patients responding to therapy in terms of a sustained virological response, however, is relatively low. One possible reason for the lack of effectiveness might be neutralization of the drug by hosts inhibitory factors. Recent kinetic studies suggested that high doses of IFN‐α, especially during the initial phase of therapy, might improve the virological response rates. Eighteen patients infected chronically with HCV were treated with IFN‐α either at a standard dose (3 × 106 to 6 × 106 IU IFN‐α three times weekly) for 6 to 12 months or with an intensified therapy (6 × 106 IU IFN‐α daily) for at least one month. As surrogate parameter for the intracellular effect of the drug, MxA gene expression was quantified in RNA preparations from peripheral blood mononuclear cells. Beta‐2‐microglobulin (β2M) concentrations were measured in serum. Serum HCV RNA titers were monitored in parallel. When compared to healthy individuals, untreated patients infected chronically with HCV were found to express 2.8‐fold higher amounts of MxA specific transcripts. MxA gene expression and serum ß2M concentrations were found to be induced after administration of IFN‐α, independent of the virological response not only during the initial phase of the intensified therapy but also over several months during standard therapy. It is concluded from these results that both early non‐effectiveness of high dose IFN‐α therapy as well as long‐term non‐effectiveness of standard therapy are not due to IFN‐α inhibitory or neutralizing elements in serum. J. Med. Virol. 62:318–326, 2000.

Clinical staging of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common cancers in the world. The resistance of HCC to existing treatments and the lack of biomarkers for early detection make it one of most hard-to-treat cancers. Surgical tumor resection, including liver transplantation, remains the only curative modality for HCC. Several clinical prognostic models have been developed for the staging of patients with HCC, but there is no general consensus which is the most reliable. There are restrictions in the use of these prognostic scores, because most scores were not validated in different countries and the HCCs differ in their biological behavior. A further problem is the heterogeneity of patients with HCC (with or without cirrhosis, compensated or decompensated cirrhosis and treated or nontreated patients); many studies combine patients with and without cirrhosis and patients with compensated and decompensated cirrhosis. The clinical efficiency of these scores is therefore limited to patients with HCC. Nowadays, DNA microarray technology has revolutionized the understanding of the molecular basis of HCC; therefore, many studies aimed to discover biomarkers for cancer staging, for prediction of recurrence, for prognosis, and for treatment selection. Most of the studies are too small for the development of predictors and at the moment microarray technology is too expensive for daily clinical use. Therefore, DNA microarray technology is at the moment not an established method in the prediction of the prognosis of patients with HCC. Future prognostic studies should include different predictors and should be performed in selected patient populations to determine whether specific prognostic indicators are more relevant at different stages of liver disease.

Hepatitis C virus virology and new treatment targets.

Hepatitis C virus (HCV) infection is the leading cause of chronic liver disease. An estimated 130 million people worldwide are persistently infected with HCV. Almost half of patients who have chronic HCV infection cannot be cured with the standard treatment consisting of pegylated IFN-α and ribavirin. For those patients who do not respond to this standard antiviral therapy, there is currently no approved treatment option available. Recent progress in structure determination of HCV proteins and development of a subgenomic replicon system enables the development of a specifically targeted antiviral therapy for hepatitis C. Many HCV-specific compounds are now under investigation in preclinical and clinical trials.

Lack of clinical evidence for involvement of hepatitis C virus interferon-α sensitivity-determining region variability in RNA-dependent protein kinase-mediated cellular antiviral responses.

The hepatitis C virus (HCV) interferon‐α (IFN‐α) sensitivity‐determining region (ISDR) has been shown to suppress double‐stranded RNA‐dependent protein kinase (PKR) activity in vitro in a yeast PKR expression system. Since variability of ISDR was shown to correlate with nonresponsiveness to IFN‐α therapy in chronically HCV‐infected patients, it has been suggested that prototype ISDR might be a viral inhibitor of cellular PKR. The present study evaluates the biological significance of ISDR variability in situ, relating it to PKR‐mediated cellular antiviral responses within the liver. ISDR variability was determined in patients chronically infected with HCV genotypes 1a, 1b, and 3a by direct sequencing using liver‐derived RNA preparations as starting material. As surrogate parameters for PKR‐mediated cellular responses, hepatic endogenous IFN‐α gene expression as well as MxA expression were analysed by a competitive, quantitative reverse transcription‐polymerase chain reaction technique. Irrespectively of intra‐ or intergenotypic ISDR amino acid substitutions, ISDR variability was found not to correlate with endogenous hepatic IFN‐α or with hepatic MxA gene expression. The data suggest that at least two prominent PKR‐mediated cellular responses might be largely unaffected by HCV ISDR variability. J. Med. Virol. 61:29–36, 2000.

Interferon‐α therapy does not modulate hepatic expression of classical type I interferon inducible genes

Hepatitis C virus (HCV) infection is a major cause of chronic liver disease. Treatment with interferon‐alpha2 (IFN‐α2) can induce viral clearance and marked biochemical and histological improvement. IFN‐α2 treatment has been shown to stimulate the expression of type I IFN regulated genes in peripheral blood mononuclear cells (PBMCs) of hepatitis C patients; however, whether it affects hepatic expression remains unknown. This study thus aimed at comparing hepatic gene expression with particular emphasis on type I IFN inducible genes in patients with chronic hepatitis C before and during an IFN‐α2 monotherapy. Responsiveness to IFN‐α2 therapy was monitored by determining serum and hepatic viral load. Differential gene expression analysis was performed by two different techniques, namely suppression subtractive hybridization (SSH) and differential display (DD). Expression of two prototype type I IFN regulated genes was quantified in further PBMC and liver samples. Among different genes found to be up‐regulated during an effective, that is, virus clearing, IFN‐α treatment, only a single one was identified which can be accounted to type I IFN responsive genes. Parallel quantitative real time PCR analyses demonstrated significant induction of the type I IFN regulated genes MxA and PKR in PBMC, but not in the liver. Taken together, while IFN‐α treatment leads to the induction of type I IFN regulated genes in PBMC, such an induction appears not to occur in the liver of hepatitis C patients. The mechanism by which IFN‐α treatment causes viral clearance might be independent of hepatic activation of type I IFN regulated genes. J. Med. Virol. 80:1912–1918, 2008.

Acute paranoid psychosis as sole clinical presentation of hepatic artery thrombosis after living donor liver transplantation

BackgroundHepatic artery thrombosis is a devastating complication after orthotopic liver transplantation often requiring revascularization or re-transplantation. It is associated with considerably increased morbidity and mortality. Acute cognitive dysfunction such as delirium or acute psychosis may occur after major surgery and may be associated with the advent of surgical complications.Case presentationHere we describe a case of hepatic artery thrombosis after living-donor liver transplantation which was not preceded by signs of liver failure but rather by an episode of acute psychosis. After re-transplantation the patient recovered without sequelae.ConclusionThis case highlights the need to remain cautious when psychiatric disorders occur in patients after liver transplantation. The diagnostic procedures should not be restricted to medical or neurological causes of psychosis alone but should also focus vascular complications related to orthotopic liver transplantation.