Elsevier

Clinica Chimica Acta

Volume 409, Issues 1–2, 3 November 2009, Pages 90-95
Clinica Chimica Acta

Impact of quality control accepted inter-laboratory variations on calculated Fibrotest/Actitest scores for the non-invasive biochemical assessment of liver fibrosis

https://doi.org/10.1016/j.cca.2009.09.005Get rights and content

Abstract

Introduction

Non-invasive, i.e. serum-based assessment of liver fibrosis is still an important diagnostic challenge although multiple single and multiparametric panels of biomarkers have been suggested.

Aim

Two approaches were followed to determine the diagnostic reliability of Fibrotest and Actitest, two commercially distributed non-invasive multiparametric tests for staging and grading of liver fibrosis.

Methods

(i) Haptoglobin, ALT, gammaGT, α-2-macroglobulin, apolipoprotein A1 and bilirubin, required for calculation of respective scores, were determined in sera of 4 patients with histologically defined stages of fibrosis (F1–F4) and activities of fibrogenesis (A1–A3). Analytes were determined by 6 quality controlled external laboratories. Inter-laboratory variations of the calculated Fibrotest score for staging and Actitest score for grading (BioPredictive™), and their error ratios referred to histologic results were calculated. (ii) The variability of respective Fibrotest/Actitest scores depending on 64 selected combinations of analytes within the accepted ranges of analyte-specific maximum/minimum limits given by the external quality control of the German Association of Clinical Chemistry and Laboratory Medicine (DGKL) was calculated.

Results

(i) Fibrotest and Actitest scores were largely reproducible among the different laboratories. However, the error ratio was 77% for all results calculated by both, Fibrotest and Actitest when referred to histologic findings. (ii) Calculated scores of stages varied between F2 (9%), F3 (31%), F3–F4 (6%), and F4 (54%) (Fibrotest), and A1/A2 (48%), A2 (9%), A2–A3 (5%), and A3 (38%) for grades of fibrogenic activity.

Conclusion

Despite reproducibility of Fibro- and Actitest scores among the six laboratories, large scale investigation displayed high levels of variability depending on inter-laboratory differences that were still in a quality controlled, analytically acceptable range. Calculated scores coincided with histologic findings in less than 25% of all cases. Thus, the diagnostic accuracy of these tests must be considered as low, if histology is accepted as reference standard.

Introduction

High prevalences of hepatitis B (HBV; ca. 300 million) and C (HCV; ca. 170 million) infections in addition to increasing numbers of non-alcoholic steatohepatitis (NASH), fatty liver disease (NAFLD), or alcoholic steatohepatitis (ASH) make chronic inflammatory liver diseases, resulting in the destruction of liver parenchyma and its replacement by scar tissue (fibrosis), a major health burden worldwide [1]. Fibrosis is characterized by the excess deposition of extracellular matrix (ECM) involving molecular and histologic re-arrangement of various types of collagens, proteoglycans, structural glycoproteins and hyaluronic acid (hyaluronan). The deposition of ECM in the space of Disse (perisinusoidal fibrosis), the generation of (incomplete) subendothelial basement membranes, and the strangulation of hepatocytes by a surrounding matrix impair not only hepatic blood flow, but also the biosynthetic function and clearance capability of hepatocytes and other cell types [2], [3]. Thus, diagnosis, follow-up and therapeutic monitoring of fibrogenesis, i.e. the active process of generation of new ECM are of great clinical importance. Currently, this is practised mostly by the invasive procedure of biopsy and consecutive histological evaluation based on numerical scoring systems such as Knodell, Ishak, METAVIR, Scheuer, Desmet and others, leading to numerical grading of necroinflammatory activity (in general the driving force of fibrogenesis) and staging, i.e. extent of fibrosis [2], [3]. However, this “gold standard” has serious drawbacks for diagnosis and follow-up [4], [5]. Therefore, the development of non-invasive, serum- or plasma-based biomarkers of fibrogenesis is an important goal, which might be reached by two, principally different approaches [6]: Class I fibrosis markers are serum analytes, which reflect ECM turnover and/or fibrogenic cell changes in the liver. They are mostly cost intensive, single laboratory tests, and hypothesis-driven, i.e. they are derived from the translation of pathogenetic mechanisms into clinical application. Class II fibrosis markers compile blood analytes based on algorithmic evaluation of commonly observed functional alterations that do not necessarily reflect ECM metabolism and/or fibrogenic cell changes. These panels of analytes are not hypothesis-driven but based on empiric observations [6], [7].

Fibrotest and Actitest scores are such class II blood tests that combine the quantitative results of six serum-based markers with the patients' age and gender in a patented algorithm in order to generate data estimating the fibrotic stage and necroinflammatory activity, respectively, corresponding to the well-established METAVIR score [8] for stages F0–F4 and grades A0–A3 [9], [10].

This study was performed to test the reliability of Fibrotest and Actitest with respect to the analytic variability still tolerated by the external quality control of Germany. Furthermore calculated Fibrotest and Actitest scores were compared with the results of the METAVIR classification system based on histology. The data point to serious limitations, which should be considered when using the investigated non-invasive tests in the follow-up of patients with chronic fibrogenic liver disease.

Section snippets

Study population and histologic staging and grading of liver fibrosis

4 randomly selected patients with HCV RNA detectable by PCR who had undergone liver biopsy between July and October 2008 were included in this study. Serum samples were taken at the time of biopsy and before any antiviral therapy. Biopsies were staged blindly according to the METAVIR group scoring system by one certified pathologist (N. G.) from the Department of Pathology, University Hospital of the RWTH Aachen as follows: grade A1 to A3 for the degree of necroinflammatory activity (A1 = mild

Theoretical calculations of Fibrotest and Actitest scores based on various combinations of minimum and maximum permissible values of the parameters required as evaluated by the external quality control system

To determine the effect of intra-analytical variations still approved by the external quality control system of the German Society of Clinical Chemistry and Laboratory Medicine (DGKL) on the calculation of Fibrotest and Actitest scores, we constructed 64 combinations with various combinations of the parameter-specific sub-maximum and sub-minimum values that were still within the accepted range of the external quality control system. These 64 combinations were submitted anonymously on the

Discussion

One can expect in the not too distant future effective therapies specifically aimed to reduce fibrogenesis, to increase fibrolysis (resolving fibrosis) or even to prevent the fibrotic organ transition [13], [14]. Although liver histology is currently considered as gold standard for diagnosis and quantification of this disease, its limitations are obvious for both, the patient and clinician [15]. Apart from its invasive nature and all associated complications that may occur, studies on sampling

Acknowledgements

We thank all laboratories listed in Table 1 for their participation in this study.

Dr. I. Hasdemir, Department of Medicine III, University Hospital of the RWTH Aachen, is thankfully acknowledged for providing serum samples of randomly selected patients with chronic liver diseases and different stages of fibrosis and Prof. Gassler, Department of Pathology, University Hospital of the RWTH Aachen, is appreciated for histological evaluation of the liver specimens.

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