January 30, 2012

Stem cells may shed light on hepatitis, MIT researchers find


Sangeeta Bhatia, MIT professor of health sciences and technology and electrical engineering and computer science

Monday, January 30, 2012

By Lori Valigra

Researchers at MIT and their colleagues said they have devised a way to produce liver-like cells from stem cells, a key step in studying why people respond differently to Hepatitis C.

An infectious disease that can cause inflammation and organ failure, Hepatitis C has different effects on different people, but no one is sure why, the researchers said in a press release from MIT. Some people are very susceptible to the infection, while others are resistant.

The researchers said that by studying liver cells from different people in the lab, they may determine how genetic differences produce these varying responses. However, liver cells are hard to get and very difficult to grow in a lab dish because they tend to lose their normal structure and function when removed from the body.

The researchers, from MIT, Rockefeller University and the Medical College of Wisconsin, have come up with a way to produce liver-like cells from induced pluripotent stem cells (iPSCs), which are made from body tissues rather than embryos. Those liver-like cells can then be infected with Hepatitis C and help scientists study the varying responses to the infection.

The scientists claim this is the first time an infection has been made in cells derived from iPSCs. Their new technique is described in the Jan. 30 issue of the Proceedings of the National Academy of Sciences. The development, they said, may also eventually enable personalized medicine, in which doctors could test the effect of different drugs on tissues derived from the patient being treated and then customize therapy for that patient.

The new study is a collaboration between Sangeeta Bhatia, professor of health sciences and technology and electrical engineering and computer science at MIT; Charles Rice, professor of virology at Rockefeller; and Stephen Duncan, professor of human and molecular genetics at the Medical College of Wisconsin.

The iPSCs are derived from normal body cells, often skin cells. By turning on certain genes in those cells, the scientists can revert them to an immature state that is identical to embryonic stem cells, which can turn into any cell type. Once the cells become pluripotent, they can be directed to become liver-like cells by turning on genes that control liver development.

The researchers’ goal is to take cells from patients who have unusual reactions to hepatitis C infection, transform them into liver cells and study their genetics to see why people respond as they do. “Hepatitis C virus causes an unusually robust infection in some people, while others are very good at clearing it. It’s not yet known why those differences exist,” Bhatia said in a statement.

Bhatia is a 2009 Mass High Tech Women to Watch honoree.


UCLA researchers developing new hepatitis C treatment


Hepatitis C virus  Credits: Robin Wulffson, MD

January 30, 2012

Robin Wulffson, M.D. LA Health Examiner

Hepatitis C is a disease that impacts approximately 160 million individuals worldwide; it can lead to liver cancer and cirrhosis. Researchers from UCLA’s Jonsson Comprehensive Cancer Center have identified a cell-permeable peptide that inhibits a hepatitis C virus protein and blocks viral replication. The finding could lead to a new treatment for the disease. Senior author Dr. Samuel French, an assistant professor of pathology and senior author of the study, and his team published their results January 30 in the peer-reviewed journal Hepatology. It builds on previous work by the French laboratory that identified two cellular proteins that are important factors in hepatitis C virus infection.

French and his team initially set out to identify the cellular factors involved in hepatitis C replication and, using mass spectrometry, found that heat shock proteins (HSPs) 40 and 70 were important for viral infection. HSP70 was previously known to be involved; however, Dr. French noted that HSP40 was linked for the first time to hepatitis C infection. The researchers further showed that the natural compound Quercetin, which inhibits the synthesis of these proteins, significantly inhibits viral infection in tissue culture.

In the study, the researchers demonstrated that the viral non-structural protein 5A (NS5A) directly binds to HSP70 and mapped the site of the NS5A/HSP70 complex on NS5A. While HSP70 was previously shown to bind NS5A in cells, a direct NS5A/HSP70 interaction and complex formation was established in this study. In an effort to stop this interaction, they tested peptides that might inhibit HSP70.

“This is important because we’ve developed a small peptide that binds to that site and blocks the interaction between the proteins that is important for viral replication,” French said. “This is another, potentially highly efficacious way to block replication of hepatitis C.”

Conventional treatments for hepatitis C, interferon and ribavirin, can have significant side effects. A new drug targeting cellular proteins rather than viral proteins would be a valuable addition to the treatment arsenal, noted Dr. French. He explained, “We were surprised that this peptide works this well. While its mechanism is different, the activity of this peptide is comparable to other newly developed anti-virals.”

The study, done in tissue culture, shows that the peptide gains entry into the cell easily and blocks the cascade of cellular events that allows the virus to replicate, French said. Blocking the HSP70 protein rather than a viral protein also reduces the chance of patients with the hepatitis C virus developing resistance to the peptide.

“There’s no direct pressure on the virus, so it is less likely to mutate and develop resistance,” Dr. French said. “The goal is to achieve a sustained response, essentially a cure, meaning there is no more virus replication. There are a lot of drugs coming out now that are designed to stop hepatitis C replication, but resistance is still an issue. About 10-20% of patients on the new drugs become resistant. This new peptide may help combat resistance.”

Going forward, Dr. French and his team are testing variants of the newly discovered peptide to see if they can develop one with an even higher affinity and can decrease the size of the peptide to improve cellular penetration and liver targeting. The new and improved peptides will be tested in animal models.

This peptide “may be a candidate for hepatitis C therapy,” the study states. “Considering the potency of the peptide in suppressing viral translation levels, treatment with this peptide may significantly improve the efficacy of conventional treatments in patients who become resistant to conventional therapies.”

The study was supported in part by the National Institutes of Health and by the California Center for Antiviral Drug Discovery at the University of California.


A multi-disciplinary approach to treating hepatitis C with interferon and ribavirin in alcohol-dependent patients with ongoing abuse

Journal of Hepatology
Volume 56, Issue 2 , Pages 334-340, February 2012

Caroline Le Lan, Anne Guillygomarc’h, Hélène Danielou, Gérard Le Dréau, Fabrice Lainé, Claude Védeilhié, Yves Deugnier, Pierre Brissot, Dominique Guyader, Romain Moirand

Received 14 March 2011; received in revised form 1 May 2011; accepted 11 May 2011. published online 13 July 2011.


Background & Aims

Guidelines recommend 6months of alcohol abstinence before treating hepatitis C (HCV). Abstinence is difficult for alcohol-dependent patients to achieve. This study evaluated HCV treatment in alcoholic patients with ongoing consumption or less than 6months of abstinence.


A multidisciplinary management model was built by a liver unit and two centers involved in the care of addict patients. Patients were included in a prospective observational study of treatment with pegylated interferon and ribavirin if they presented alcohol dependence with ongoing intoxication or abstinence of less than 6months. Pre-therapeutic evaluation and follow-up were multidisciplinary, and addiction care was personalized to patient condition and willingness. Alcohol abstinence or reduction was encouraged but not mandatory. The primary end point was sustained virological response (SVR). Results were compared to a control group of patients matched for genotype, viral load, fibrosis stage, sex, and age.


A total of 73 patients treated between 2002 and 2008 were included in the study. Intent to treat analysis showed an SVR in 48% (35/73) of patients versus 49% (36/73) of controls. Low viral load and length of abstinence during treatment were independently associated with SVR. During treatment, 20 (27%) patients were abstinent, 23 (32%) had controlled consumption, and 24 (33%) had excessive consumption. At the end of the follow-up, 22 (30%) patients were durably abstinent.


A multidisciplinary approach allowed HCV treatment in alcohol-dependent patients with a satisfactory SVR rate and positive effects on addiction behavior.


HIV’s Increasingly Dangerous Sidekick


30 January 2012

If you are HIV-positive you have a lot to deal with on your plate already. The last thing you need is to have your liver ruined by a Hepatitis C infection as well. Unfortunately it appears that the co-infection rate is soaring, according to the latest National AIDS Trust (NAT) report.

Recent studies have shown that the amount of gay men discovering that they are HIV-positive is increasing at a dramatic rate, especially in the younger population. Every week there seems to be a new statistic telling us that we are going to catch it, that every potential partner is going to infect us with it, and as a result we have started to moderate our behaviour to allow for this. However, there is another statistic which is increasing alongside this; infection of Hepatitis C is increasing too. According to the NAT’s Report, 9% of HIV-positive people in the UK are infected with Hepatitis C too, which doesn’t sound like a lot. If you factor in statistics like the amount of estimated people who are positive, and the increases in young men who have been infected, you end up looking at scary numbers. Deborah Jack, Chief Executive of NAT, told So So Gay, ‘The rate of HIV-positive gay men co-infected with hepatitis C in the UK is too high. It is crucial for this to be addressed as a strategic priority in gay men’s health promotion.’

According to the report, those who test positive for Hepatitis C are marginalised and vilified by the HIV-positive community in the same way that those with HIV have been by the gay community at large. One participant quoted in the report said, ‘Hep C is not yet owned by the gay community like HIV, and if it isn’t owned, then it is outside and more stigmatised… Even within the gay community, and the HIV community too, it has created a ‘them’ and ‘us’ type situation’. This seems ridiculous for two main reasons: one of the biggest things we hear from the HIV community is how they wish to be treated normally, and this paints a very hypocritical picture; and unlike HIV, Hepatitis is curable in the some cases.

The report goes on to discuss prevention and treatment of Hepatitis C, quoting a success rate of up to 80% early treatment rate in HIV-positive individuals. The key it seems, as per usual, is in the power of knowledge: knowing about the risks and how to avoid risky situations is what differs those who have it and those who don’t.


More must be done to reduce hepatitis C transmission amongst HIV positive gay men

Jan. 27, 2012

More must be done to reduce hepatitis C transmission amongst HIV positive gay men

NAT has launched a report, ‘HIV and Hepatitis C Co-infection’, which looks at hepatitis C co-infection among HIV positive gay men and the UK’s response to this growing health challenge. Like HIV, hepatitis C* is a blood-borne virus and approximately 9% of HIV positive people in the UK also have hepatitis C. Having both conditions at the same time can have severe health implications, with liver disease caused by hepatitis B and/or C a leading cause of serious illness and death in people with HIV.

7% of HIV positive gay men are co-infected with hepatitis C. Of those who successfully clear hepatitis C through treatment, a significant percentage get re-infected within a short time. Infections among gay men are largely due to sexual risk factors, thought to include unprotected anal sex, fisting, use of sex toys, group sex – though drug use may also have a role. There have been some important information campaigns in the gay media around the risk of hepatitis C for HIV positive gay men but -

  • There is no explicit national strategic approach to tackling this issue
  • Hepatitis C testing recommendations for gay men at risk are not being adequately implemented
  • A wide range of possible sexual risk factors are cited which can detract from focussing on what are clearly higher risk behaviours
  • Stigma around hepatitis C in the gay community and amongst people with HIV hampers prevention efforts and harms gay men living with hepatitis C

Deborah Jack, Chief Executive of NAT (National AIDS Trust), comments:

‘The rate of HIV-positive gay men co-infected with hepatitis C in the UK is too high. It is crucial for this to be addressed as a strategic priority in gay men's health promotion.

‘It is vitally important that, as recommended, all people diagnosed with HIV are annually screened for hepatitis C infection and this should be made a requirement in the commissioning of all relevant services (in a recent audit only 66% had had an annual test). Clinics and health promoters need to provide intensive advice and support to gay men at significant risk of hepatitis C transmission. To that end, consensus is urgently needed on the key risk factors for sexual transmission so clear and appropriate recommendations can then be made.

‘We also strongly urge gay men not to rely on their sexual partners’ disclosure of their HIV or hepatitis C status as a high proportion are unaware they are infected, which is certainly fuelling onward transmission. And even when diagnosed, disclosure can be difficult - we need to start challenging hepatitis C stigma as well as HIV stigma - both are unfair, ill-informed and destructive.’

- Ends -

Notes to the editor:

* Hepatitis C is a blood-borne virus that can cause liver inflammation and other long-term health problems. Unlike hepatitis A and hepatitis B there is no vaccine against hepatitis C.

Approximately 25% of people naturally clear the infection at the acute phase (the first six months of infection), but for the remaining 75%, hepatitis C infection becomes chronic. Treatment for hepatitis C is available and response rates vary from approximately 40% to 80% and therapy can cause significant side-effects. Unless successfully treated, the infection can lead to liver fibrosis and cirrhosis, end stage liver disease and liver cancer, and death.

The full report can be downloaded here.

For further information please contact:
Charli Scouller
Communications Manager
020 7814 6733


NAT (National AIDS Trust) is the UK’s leading charity dedicated to transforming society’s response to HIV. We provide fresh thinking, expertise and practical resources. We champion the rights of people living with HIV and campaign for change.

Shaping attitudes. Challenging injustice. Changing lives.


Hepatitis C Virus Studied as Risk Factor in Liver Cancer

Infectious Disease Special Edition ISSUE: JANUARY 2012 | VOLUME: 1

by George Ochoa

Two Mayo Clinic studies have clarified the importance of chronic hepatitis C virus (HCV) in the rising trend of liver cancer, or hepatocellular carcinoma (HCC).

One study (Yang et al. Mayo Clin Proc. 2012;87:9-16) analyzed longitudinal trends in the incidence, etiology, treatment of HCC and survival in community residents in Olmsted County, Minn. The researchers found that in earlier periods (1976-1990, 1991-2000), alcohol use was the most common risk factor, but, in 2001 to 2008, HCV filled that role. At the same time, HCC incidence rose dramatically, from 3.5 per 100,000 person-years for the 1976 to 1990 time period, to 3.8 for the period from 1991 to 2000, to 6.9 for the 2001 to 2008 period.

Study author W. Ray Kim, MD, associate professor of medicine, Mayo Clinic College of Medicine, Rochester, Minn., wrote in an email: “The biggest and unique risk factor of HCC is underlying liver disease. Our data showed that previously alcohol, more recently HCV has become the underlying cause.”

The second study (Shire et al. Mayo Clin Proc. 2012;87:17-24) investigated a sample of Somali immigrants seen at Mayo Clinic from July 1, 1996, to Oct. 31, 2009. Non-Somali Olmsted County residents served as controls. The frequencies of chronic hepatitis B virus (HBV) and HCV, and their associations with HCC, were studied. Both HBV and HCV occurred frequently in the sample of Somalis, but HCV was the major risk factor for HCC. There were significant differences in the HCV genotype distributions between Somalis and non-Somalis.

The high prevalence of HCV in the Somali sample came as a surprise to the researchers. “We didn’t expect it,” Abdirashid M. Shire, PhD, assistant professor of medicine, Division of Gastroenterology and Hepatology, Mayo Clinic College of Medicine, said in an interview. The study, Dr. Shire added, “supports the value of early detection” of HCV. “It’s very important to screen for HCV.” Dr. Kim reported that his study also “indirectly” supports the value of early detection and treatment to improve outcomes.

—Drs. Kim and Shire had no relevant financial disclosures.


Muscle Wasting Is Associated With Mortality in Patients With Cirrhosis

Dr. Aldo J. Montano-Loza discusses his manuscript "Muscle Wasting Is Associated With Mortality in Patients With Cirrhosis" To view the print version of this abstract go to http://bit.ly/yAAXok.

Blood Test Instead of Biopsy Identifies Liver Damage


Siemens is marketing the first rapid, automated biomarker test for diagnosing and assessing liver fibrosis.

The ELF test (Enhanced Liver Fibrosis test) takes approximately one hour to complete and requires only a blood sample. The process is therefore less invasive but just as reliable as the previously required biopsy, and it usually takes about a week to deliver a biopsy result.

The new test was developed by Siemens Healthcare in collaboration with University College London and can be used as a routine test on the Siemens ADVIA Centaur Immunoassay System.

Liver fibrosis is the result of chronic liver damage caused by vi-ral hepatitis, alcoholic cirrhosis, or fatty liver disease. It is characterized by scarring of the liver tissue, which can lead to cirrhosis or cancer of the liver over the long term — a frequent cause of death worldwide.

At present, the “gold standard” for assessing the severity of a liver fibrosis is a liver biopsy, which involves the removal of a small amount of liver tissue. This biopsy has drawbacks, however: It is painful; it entails some risk for the patient; and it tests only a small sample of the liver.

With the automated ADVIA Centaur ELF Test, a fast and mini-mally invasive technique is now available for determining both the severity of a liver fibrosis and the risk that it will worsen. The test examines three direct blood serum biomarkers: hyaluronic acid (HA), procollagen III N-terminal propeptide (PIIINP), and the tissue inhibitor of metalloproteinase 1 (TIMP-1). These direct biomarkers are molecules that are involved in the formation of fibrosis. In the test, special reagents react with the biomarkers and generate light in the process. The greater the intensity of this chemiluminescence, the greater the presence of the biomarker.

A special algorithm is used to convert the results of the three biomarkers into the ELF score, which indicates the degree of fibrosis. The combination of three biomarkers increases the accuracy of the test. As an international clinical trial has shown, the ELF test can precisely differentiate between slight, moderate, and serious cases of fibrosis. In the event of slight or moderate fibrosis, patients normally have no symptoms. Doctors can thus intervene before significant damage to the liver and monitor the progress of therapy.

To complement the ELF test, Siemens is also offering imaging and laboratory diagnostic technologies like hepatitis blood tests and ultrasound systems that help physicians identify liver fibrosis at an early stage and monitor its development.

Dr. Norbert Aschenbrenner | Source: Siemens InnovationNews
Further information: www.siemens.com/innovationnews


The ELF Program

In 1997 a group of European investigators convened by Professor Michael Arthur embarked on a project led by Professor William Rosenberg funded by Bayer Healthcare to identify serum markers of liver fibrosis. This program of research has been continuously active for over a decade and has resulted in the identification of a panel of direct markers that have been validated, CE marked, and are now being marketed in Europe by iQur Limited and Siemens.


The original European Liver Fibrosis project recruited over 1,000 patients having investigation with a liver biopsy at 13 centres across Europe. The subjects had a wide range of CLD reflecting clinical practice. Over 40% had CHC or CHB. All patients had a fasting serum sample taken at the time of their biopsy. This was sent to a central laboratory for analysis of a panel of candidate analytes representing direct markers of fibrosis as well as a number of indirect makers. The individual sandwich ELISA test for each of the direct markers were carefully developed to a high standard of accuracy, reproducibility and repeatability. Using logistic regression and multivariate analysis those markers that most accurately reflected the stage of liver fibrosis assigned by a central pathologist were identified in a “training” cohort of 521 patients and then confirmed in a “validation” cohort of 400. This revealed that the combination of HA, P3NP and TIMP-1 combined in an algorithm, originally incorporating Age, could be used to determine the severity of liver fibrosis with good accuracy.14 Subsequently the team have established that Age could be omitted from the algorithm to generate the Enhanced Liver Fibrosis test or ELF Test.

The original European Liver Fibrosis project recruited over 1,000 patients having investigation with a liver biopsy at 13 centres across Europe. The subjects had a wide range of CLD reflecting clinical practice. Over 40% had CHC or CHB. All patients had a fasting serum sample taken at the time of their biopsy. This was sent to a central laboratory for analysis of a panel of candidate analytes representing direct markers of fibrosis as well as a number of indirect makers. The individual sandwich ELISA test for each of the direct markers were carefully developed to a high standard of accuracy, reproducibility and repeatability. Using logistic regression and multivariate analysis those markers that most accurately reflected the stage of liver fibrosis assigned by a central pathologist were identified in a “training” cohort of 521 patients and then confirmed in a “validation” cohort of 400. This revealed that the combination of HA, P3NP and TIMP-1 combined in an algorithm, originally incorporating Age, could be used to determine the severity of liver fibrosis with good accuracy.14 Subsequently the team have established that Age could be omitted from the algorithm to generate the Enhanced Liver Fibrosis test or ELF Test.

External Validation

Subsequent to the original study, investigators around the world have conducted validation studies in independent populations to further assess the performance of the markers. Studies in CHC, NAFLD and PBC have all confirmed that the markers accurately reflect the severity of fibrosis as staged on liver biopsy 15-19. In these studies the area under the receiver operator characteristic curve (AUROC) ranges around 0.8. This level of performance is considered to be a threshold for acceptance in clinical practice.

Longitudinal follow-up

However the investigators have been limited in their ability to assess the performance of the markers by the errors inherent in biopsy staging. In an attempt to overcome this “glass ceiling” the ELF team have begun to investigate the ability of the serum markers to predict long-term clinical outcomes of CLD including the development of portal hypertension, decompensation of CLD, the development of hepatocellular cancer, liver transplantation and death from liver disease, as well as all cause mortality. Whilst not yet complete, interim analysis of the 7 year follow-up of over 500 patients has shown that the ELF markers are at least as good, if not better than liver histology at predicting clinical outcome. Similar work has been conducted in a cohort of patients with PBC. 20

Advantages over single markers

Systematic reviews of the literature have consistently shown that single markers of fibrosis can detect cirrhosis with some degree of accuracy. However single marker test are less accurate than panels of markers in detecting lesser degrees of fibrosis 21,22. While the detection of cirrhosis is important, clinical judgement can often identify patients who are likely to have end-stage CLD. It is of greater importance to be able to detect patients with mild or moderate fibrosis which is usually asymptomatic, in order to be able to intervene with life-style modification or treatment before the liver becomes irreparably damaged. It is in the screening and management of patients with CLD that the ELF markers have great potential.

Advantages compared to indirect markers

The careful and extensive validation program employed by the ELF development team has taken years to come to fruition. A number of algorithms have been derived employing indirect markers that are available in general clinical chemistry laboratories as routine analytes. While the performance of many of these algorithms such as Fibrotest/Fibrosure, APRI, Forns Index and Hepascore is good, the majority are unreliable in patients undergoing treatment for viral hepatitis (where aminotransferases are altered due to therapy) or when bilirubin levels may be elevated due to haemolysis (such as when Ribavirin is administered) or cholestasis. 23-29

Use in clinical practice and Impact on patient management

The coming year will see the introduction of the ELF test into clinical practice and its impact in the management of patients will begin to be appreciated. Doctors in primary care are flooded with patients with obesity and hazardous drinking all of whom could be at risk of CLD. The ability to use a simple blood test to accurately identify those with significant liver disease will greatly aid triage and the appropriate targeting of interventions including weight loss, exercise and drug interventions. In secondary care the ELF test will be of great use in the early evaluation of patients with a wide range of CLD. It will not replace liver biopsy in the detailed assessment of liver inflammation, architectural damage of pathology. However it can be used to prioritise patients for investigation, to determine the severity of fibrosis in patients unwilling or unable to undergo biopsy and to compliment biopsy given the inaccuracies that surround histological staging arising from sampling error and observer error. Once a diagnosis of CLD has been made most patients will undergo some form of treatment and long-term follow-up. Further repeated biopsies are not acceptable in the vast majority of patients but further knowledge of the severity of fibrosis, its progression or regression would be highly valuable to both the patient and the doctor. The ELF test will make this information accessible through a simple blood test that could be repeated at frequent intervals.


Figure 1 AUROC curve of ELF predicting stages 0,1 Vs 2-6 in NAFLD cohort (none or mild fibrosis from significant fibrosis Ishak classification)

Future developments

While the introduction of the ELF test represents a major advance in the diagnostic armoury available clinician screening for CLD managing CLD further challenges remain. The ELF markers have been developed for a range of CLD. In specific diseases other combinations of markers may perform better. So far the development of algorithms such as ELF has relied on the testing of candidate markers. New discovery approaches such as metabonomics, proteomics and transcriptomics that make no assumptions about the relevance of specific molecules to disease processes may uncover new markers that complement or enhance the existing panels. Imaging of the liver using a variety of modalities including ultrasound, elastography and magnetic resonance imaging has advanced in parallel with non-invasive marker testing 30,31. The integration of these complementary modalities should further increase the diagnostic power available to clinicians. Determining the optimal combinations and suitability for specific CLD will be challenging.

The discovery of the ELF markers represents the dawn of a new era of early detection of treatable liver disease and the effective monitoring of CLD to evaluate the impact of interventions and the course of disease.

Disadvantages of liver biopsy
  • Morbidity & mortality
  • Sampling error
  • Ordinal categorical variable to assess continuous biological process
  • Costly & time consuming- requires hospital visit and expert
  • Inter-observer variability- kappa scores 0.4-0.6
  • Error rate –up to 25-35% of ≥1 stage
  • Less experienced pathologist perform less well than “expert”
  • Cannot perform repeat biopsy at short intervals to assess liver disease or effects of therapeutics due to the hazards.

Liver Fibrosis
Overview Page
What is liver fibrosis?
Non-invasive markers of liver fibrosis

Part of the article “Biomarkers of liver disease: the enhanced liver fibrosis test”
As published in CLI October 2007

The authors
William Rosenberg MD, D.Phil
Professor of Hepatology,
The Liver Group,
University of Southampton,
Southampton, UK

Julie Parkes MD
Public Health Science & Medical Statistics,
University of Southampton,
Southampton, UK

Reference List

(14) Rosenberg WM, Voelker M, Thiel R, Becka M, Burt A, Schuppan D et al. Serum markers detect the presence of liver fibrosis: a cohort study. Gastroenterology 2004; 127(6):1704-1713.

(15) Parkes J, Bialek SR.,Bell BP ,Terrault N., Zaman A., Sofair A., Guha IN, Cross R, Harris S, Roderick PJ, Rosenberg WMC European Liver fibrosis markers accurately distinguish fibrosis severity in a cohort of patients with Chronic Hepatitis C; an external validation study. Hepatology 44[Suppl 1]. 2006.

(16) Parkes J, Cross R, Harris S, Ryder S, Irving W, Zaitoun A, The Trent Hepatitis C Research Group, Rosenberg WMC. European liver fibrosis markers accurately distinguish fibrosis severity in Chronic Hepatitis C. Journal of Hepatology. 40 [Suppl 1]. 2005.

(17) IN Guha, GP Aithal, J Parkes , PJ Roderick, S Harris , R Cross, P Kaye, SD Ryder, WM Rosenberg European liver fibrosis markers in Non Alcoholic Fatty Liver Disease (NAFLD); an external validation study. Hepatology 44 Suppl 1. 2006.

(18) J. Parkes, M. Mayo , R.Cross , S.Harris, P.J.Roderick B. Coombs, B. Huet The PUMPS Investigators and W.M. Rosenberg . European liver fibrosis markers accurately distinguish fibrosis severity in primary biliary cirrhosis; an external validation study. Hepatology 44 Suppl 1. 2006.

(19) Non-invasive markers of fibrosis in non-alcoholic fatty liver disease: validatingthe European Liver Fibrosis panel and exploring simple markers. IN Guha, J Parkes ,P Roderick , D Chattopadhyay, R Cross, S Harris, P Kaye , AD Burt, SD Ryder, GP Aithal , CP Day, WM Rosenberg. Hepatology 2007 (in press)

(20) Mayo M., Parkes J, Huet B, Combes B, Mills S., Markin R. et al. Serum fibrosis markers predict future clinical decompensation in primary biliary cirrhosis bettter than liver biopsy, bilirubin, or Mayo risk score. Hepatology 44 Suppl 1. 2006.

(21) Guha IN, Parkes J, Roderick PR, Harris S, Rosenberg WM. Non-invasive markers associated with liver fibrosis in non-alcoholic fatty liver disease 2. GUT 2006; 55(11):1650-1660.

(22) Parkes J, Guha IN, Roderick P, Rosenberg W. Performance of serum marker panels for liver fibrosis in chronic hepatitis C. Journal of Hepatology 2006; 44:462-474.

(23) Imbert-Bismut F, Ratziu V, Pieroni L, Charlotte F, Benhamou Y, Poynard T. Biochemical markers of liver fibrosis in patients with hepatitis C virus infection: a prospective study. Lancet 2001; 357(9262):1069-1075.

(24) Ratziu V, Massard J, Charlotte F, Messous D, Imbert-Bismut F, Bonyhay L et al. Diagnostic value of biochemical markers (Fibro Test-FibroSURE) for the prediction of liver fibrosis in patients with non-alcholic fatty liver disease. BMC Gastroenterology 2006; 6(6):1-13.

(25) Wai CT, Greenson JK, Fontana RJ, Kalbfleisch JD, Marrero JA, Conjeevaram HS et al. A simple noninvasive index can predict both significant fibrosis and cirrhosis in patients with chronic hepatitis C.. Hepatology 2003; 38(2):518-526.

(26) Forns X, Ampurdanes S, Llovet JM, Aponte J, Quinto L, Martinez-Bauer E et al. Identification of chronic hepatitis C patients without hepatic fibrosis by a simple predictive model.. Hepatology 2002; 36(4 Pt 1):986-992.

(27) Patel K, Muir AJ, McHutchison JG. Validation of a simple predictive model for the identification of mild hepatic fibrosis in chronic hepatitis C patients.. Hepatology 2003; 37(5):1222-1223.

(28) Hepascore: An Accurate Validated Predictor of Liver Fibrosis in Chronic Hepatitis C Infection Adams,L.A.; Bulsara,M.; Rossi,E.; de Boer,B.; Speers,D.; George,J.; Kench,J.; Farrell G; McCaughan,G.W.; Jeffrey,G.P. Clinical Chemistry 51 (10) 1-7

(29) Cales P, Oberti F, Michalak S, Hubert-Fouchard I, Rousselet MC, Konate A et al. A novel panel of blood markers to assess the degree of liver fibrosis 10. Hepatology 2005; 42(6):1373-1381.

(30) Ganne-Carríe N, Ziol M, Ledinghen V, Douvin C, Marcellin P, Castera L et al. Accuracy of Liver Stiffness Measurement for the Diagnosis of Cirrhosis in Patients With Chronic Liver Disease. Hepatology 2006; 44:1511-1517.

(31) Foucher J ,Chanteloup E,Vergniol J,Castera L,Le Bail B,Adhoute X,Bertet J,Couzigou P,de Ledinghen V. Diagnosis of cirrhosis by transient elastography (FibroScan): a prospective study. GUT 2006; 55(3):403-408.

* The ELF test is only available outside the US