Liver stiffness measurement (fibroscan®)

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elastography, fibroscan, liver stiffness measurement.

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  • Liver stiffness values are expressed in kilopascalsRange from 2.5 – 75 kPaValues around 5.5kPa were recently shown to reflect normality
  • Normal portal triads with no signs of fibrosis (stage F0)Portal fibrous expansion (stage F1)Thin fibrous septa emanating from portal triads (stage F2) Fibrous septa bridging portal triads and central veins (stage F3)Cirrhosis (stage F4), which appears as nodules of liver parenchyma separated by thick fibrous bands.
  • The experience accumulated so far suggests that the performance of LSM is optimal for advanced stages of fibrosis (ie, >F3 according to the METAVIR scoring system).This is likely due to the proportionally reduced presence of necrosis and inflammation, which are known to affect LSM. On the contrary, within intermediate stages of fibrosis the occurrence of inflammation, steatosis, cholestasis and passive/active congestion of the liver, may affect LSM.Therefore, patients with advanced fibrosis or compensated cirrhosis represent an optimal setting for the use of LSM. The ideal candidate for TE is a lean patient with severe fibrosis.
  • Additional advantage of Fibroscan is that it provides a wide range of stiffness values within the group of cirrhotic livers that would overcome one major limitation of the biopsy (i.e. one histological stage for all types of cirrhosis), and could thus provide an additional prognostic value within this group (i.e. one histological stage for all types of cirrhosis), and could thus provide an additional prognosticvalue within this group.Background: Transient elastography (FibroScan) is a new, non-invasive, rapid, and reproducible method allowing evaluation of liver fibrosis by measurement of liver stiffness. In cirrhotic patients, liver stiffness measurements range from 12.5 to 75.5 kPa. However, the clinical relevance of these values is unknown. The aim of this prospective study was to evaluate the accuracy of liver stiffness measurement for the detection of cirrhosis in patients with chronic liver disease.Methods: A total of 711 patients with chronic liver disease were studied. Aetiologies of chronic liver diseases were hepatitis C virus or hepatitis B virus infection, alcohol, non-alcoholic steatohepatitis, other, or a combination of the above aetiologies. Liver fibrosis was evaluated according to the METAVIR score.Results: Stiffness was significantly correlated with fibrosis stage (r = 0.73, p,0.0001). Areas under the receiver operating characteristic curve (95% confidence interval) were 0.80 (0.75–0.84) for patients with significant fibrosis (F.2), 0.90 (0.86–0.93) for patients with severe fibrosis (F3), and 0.96 (0.94–0.98) for patients with cirrhosis. Using a cut off value of 17.6 kPa, patients with cirrhosis were detected with a positive predictive value and a negative predictive value (NPV) of 90%. Liver stiffness was significantly correlated with clinical, biological, and morphological parameters of liver disease. With an NPV > 90%, the cut off values for the presence of oesophagealvarices stage 2/3, cirrhosis Child-Pugh B or C, past history of ascites, hepatocellular carcinoma, and oesophageal bleeding were 27.5, 37.5, 49.1, 53.7, and62.7 kPa, respectively.Conclusion: Transient elastography is a promising non-invasive method for detection of cirrhosis in patients with chronic liver disease. Its use for the follow up and management of these patients could be of great interest and should be evaluated further.Retrospective study of 711 patients with chronic liver diseases (95 with histologically proven cirrhosis).These preliminary findings need to be confirmed in long-term prospective follow-up studies to see whether liver stiffness values can predict the occurrence of clinical events in patients with compensated cirrhosis.
  • Hazard ratio as compared to LSM<10 kPa≤LSM 10.1-15 kPa 16.7 (95% CI, 3.71-75.2; P<0.001 LSM 15.1-20 kPa 20.9 (95% CI, 4.43-98.8; P<0.001LSM 20.1-25 kPa 25.6 (95%CI, 5.21-126.1; P < 0.001LSM >25 kPa 45.5 (95% CI, 9.75-212.3; P < 0.001Liver stiffness, noninvasively measured by transient elastography, correlates well with liver fibrosis stage. The aim of this prospective study was to evaluate the liver stiffness measurement (LSM) as a predictor of hepatocellular carcinoma (HCC) development among patients with chronic hepatitis C. Between December 2004 and June 2005, a total of 984 HCV-RNApositive patients, without HCC or a past history of it, visited the University of Tokyo Hospital. LSM was performed successfully in 866 patients, who gave informed consent. During the follow-up period (mean, 3.0 years), HCC developed in 77 patients (2.9% per 1 person-year). The cumulative incidence rates of HCC at 1, 2, and 3 years were 2.4%, 6.0%, and 8.9%, respectively. Adjusting for other significant factors for HCC development, patients with higher LSM were revealed to be at a significantly higher risk, with a hazard ratio, as compared to LSM<10 kPa, of 16.7 (95% confidence interval [CI], 3.71-75.2; P<0.001) when LSM 10.1-15 kPa, 20.9 (95% CI, 4.43-98.8; P<0.001) when LSM 15.1-20 kPa, 25.6 (95%CI, 5.21-126.1; P < 0.001) when LSM 20.1-25 kPa, and 45.5 (95% CI, 9.75-212.3; P < 0.001) when LSM >25 kPa. Conclusions: This prospective study has shown the association between LSM and the risk of HCC development in patients with hepatitis C. The utility of LSM is not limited to a surrogate for liver biopsy but can be applied as an indicator of the wide range of the risk of HCC development.
  • In a study of 10 000 liver biopsies, Bedossa demonstrated that the stage of fibrosis was correctly diagnosed in 65% of biopsies with length of 15 mm 75% of biopsies with length of 25 mmBedossa P et al. Hepatol 2003 ; 38 : 1449 – 57.
  • Waist circumference 80 cm in women or 94 cm in men. Specific probe is developed for obese patients.
  • Feasibility of liver transient elastography with FibroScans using a new probe for obese patientsde Lédinghen V et al. Liver International 2010 ; : 1043 – 1048. Background & Aims: Liver stiffness measurement (LSM) failure when using transient elastography occurs in 2–10% of patients, and is generally related to obesity. The aim of this prospective study was to assess the feasibility of LSM when using a new XL probe on patients with a body mass index(BMI)Z30 kg/m2. Methods: For each patient, LSM was performed using both M probe (currently available and dedicated to patients with standardmorphology) and XL probe (dedicated to overweighed patients). A blood sample was taken to assess usual biological variables and simple readily available fibrosis blood tests. Results: Ninety-nine patients were included (27 men, mean age 52 years, mean BMI 40.5 kg/m2). LSM was successful (10 valid measurements) in 45% of the cases with the M probe, vs 76% of the cases with the XL probe (P<0.001). Fifty-nine percent of those who could not be measured (< 10 valid measurements) using the M probe could successfully be measured using the XL probe. In the 44 patients successfullymeasured with both probes, LSM was correlated with the platelet count, prothrombin time, g-glutamyltransferase, aspartateaminotransferase, fasting glucose, AST platelet ratio index, Forns score and FIB-4. Conclusion: The new XL probe allows providing a higher rate of LSM than the M probe in patients with an increased BMI and shows promising results for the evaluation of liver fibrosis.
  • When interpreting LSMs, the clinical question should be defined and LSM should be interpreted in the context of a defined diagnosis, biochemical data and appropriate cutoff points.Conclusion: LSM does not represent a reliable instrument to detect the presence of advanced fibrosis and cirrhosis in patients presenting with a clinical picture of acute hepatitis.
  • When interpreting LSMs, the clinical question should be defined and LSM should be interpreted in the context of a defined diagnosis, biochemical data and appropriate cutoff points.Conclusion: LSM does not represent a reliable instrument to detect the presence of advanced fibrosis and cirrhosis in patients presenting with a clinical picture of acute hepatitis.
  • Transient elastography (FibroScan [FS]) is a novel non-invasive tool to assess liver fibrosis/ cirrhosis. However, it remains to be determined if other liver diseases such as extrahepaticcholestasis interfere with fibrosis assessment because liver stiffness is indirectly measured by the propagation velocity of an ultrasound wave within the liver. In this study, we measured liver stiffness immediately before endoscopic retrograde cholangiopancreatography and 3 to 12 days after successful biliary drainage in patients with extrahepaticcholestasis mostly due to neoplastic invasion of the biliary tree. Initially elevated liver stiffness decreased in 13 of 15 patients after intervention, in 10 of them markedly. In three patients, liver stiffness was elevated to a degree that suggested advanced liver cirrhosis (mean, 15.2 kPa). Successfuldrainage led to a drop of bilirubin by 2.8 to 9.8 mg/dL whereas liver stiffness almost normalized (mean, 7.1 kPa). In all patients with successful biliary drainage, the decrease of liver stiffness highly correlated with decreasing bilirubin (Spearman’s 0.67, P < 0.05) with a mean decrease of liver stiffness of 1.2± 0.56 kPa per 1 g/dLbilirubin. Two patients,in whom liver stiffness did not decrease despite successful biliary drainage, had advanced liver cirrhosis and multiple liver metastases, respectively. The relationship between extrahepaticcholestasis and liver stiffness was reproduced in an animal model of bile duct ligation in landrace pigs where liver stiffness increased from 4.6 kPa to 8.8 kPa during 120 minutes of bile duct ligation and decreased to 6.1 kPa within 30 minutes after decompression.Conclusion:Extrahepaticcholestasis increases liver stiffness irrespective of fibrosis. Once extrahepaticcholestasis is excluded (e.g., by liver imaging and laboratory parameters) transient elastography is a valuable tool to assess liver fibrosis in chronic liver diseases.
  • Conclusion:Extrahepaticcholestasis increases liver stiffness irrespective of fibrosis. Once extrahepaticcholestasis is excluded (e.g., by liver imaging and laboratory parameters) transient elastography is a valuable tool to assess liver fibrosis in chronic liver diseases.
  • Median weight loss of 3.0 kgLS is a direct function of central venous pressure which should be considered when assessing the degree of fibrosis.The presence of enlarged liver and/or hepato-jugular reflux on physical examination together with dilated hepatic veins and inferior vena cava, suggestive of chronic heart failure, would have helped to anticipate the findings of liverbiopsy.
  • Liver stiffness measurement (fibroscan®)

    1. 1. Liver stiffness measurement (Fibroscan®) Principles - indications - results - limitations Samir Haffar M.D. Assistant Professor of Gastroenterology
    2. 2. Fibrose Blood markers FibroScan® Imaging (US, MRI, endoscopy) Hepatic biopsy Biological work-up Clinical Examination
    3. 3. Ideal non-invasive test for diagnosis of liver fibrosis • Simple • Reproducible • Readily available • Less expensive than biopsy • Predicts full spectrum of fibrosis • Reflects changes occurring with therapy
    4. 4. Evaluation of chronic liver injury according to health care level Physical examination Liver function tests Serum Hyaluronate APRI or other simple tests Primary health care Ultrasound Fibroscan® Serum markers & algorithms Secondary health care Fibroscan® ARFI* MR elastography* Tertiary health care Liver biopsy HVPG * Promising but currently under investigation ARFI: Acoustic Radiation Force Impulse Imaging HVPG: Hepatic Venous Pressure Gradient Castéra L et al. Gut 2010 ; 59 : 861 – 866.
    5. 5. Liver stiffness • Assessed by US (FibroScan®) & more recently by MRI • Evaluates velocity of propagation of a shock wave within liver tissue (examines a physical parameter of liver tissue which is related to its elasticity) • Rationale Normal liver is viscous Not favorable to wave propagation Fibrosis increases hardness of tissue Favors more rapid propagation Bedossa P. Liver Int 2009 ; 29 (s1): 19 – 22.
    6. 6. Fibroscan® device • Electronic platform – Ultrasonic signals acquisition – Numerical signal processing • Integrated computer – Stiffness measurement – Examinations database • Dedicated probes with unique technology Vibrator (50 Hz) US Transducer (3,5 MHz) Fibroscan® (Echosens, Paris, France)
    7. 7. Position of probe & explored volume Cylinder of 1 cm wide & 4 cm long From 25 mm to 65 mm below skin surface This volume is at least 100 times bigger than a biopsy sample
    8. 8. Results Stiffness (kPa) Median value of 10 shots 3.9 Kilo Pascals  At least 10 shots  Success Rate: ≥ 60%  IQR * (kPa) Interval around median Contains 50% of valid shots ≤ 25% of median value
    9. 9. Manufacturer’s criteria for LSM interpretation • First step Number of shots ≥ 10 • Second step Success rate ≥ 60 % • Third step Interquantile range(IQR) ≤ 25% Failure Zero valid shot Unreliable results < 10 valid shots Success rate ≤ 60% IQR ≥ 25%
    10. 10. Liver stiffness values in healthy subjects 429 subjects Roulot D et al. J Hepatol 2008 ; 48 : 606 – 613. 5.2 1.5 kPa 5.8 1.5 kPa p = 0.0002
    11. 11. Liver stiffness values in healthy subjects with & without metabolic syndrome Roulot D et al. J Hepatol 2008; 48 : 606 – 613. 5.3 1.5 kPa 6.5 1.6 kPa p < 0.0001
    12. 12. Liver stiffness cut-offs in chronic liver diseases F2 Sign F3 Severe F4 Cirrhosis Matavir F0-F1 MildFibrosis Castéra L et al. J Hepatol 2008 ; 48 : 835 – 847. LSM 2.5 – 7 kPa → Mild or absent fibrosis is likely LSM > 12.5 kPa → Cirrhosis is likely
    13. 13. Progression of fibrosis in viral hepatitis Photomicrographs (magnification 40; trichrome stains) Faria SC et al. RadioGraphics 2009 ; 29 : 1615 – 1635. F 0 F 1 F 2 F 3 F 4
    14. 14. Perform LSM ≤ 6 kPa No significant fibrosis No biopsy F0 F1F Intermediate values Grey area Biopsy if results influence management F2 Implementation of other NI tests ≥ 12 kPa Advanced fibrosis No biopsy F4 Treatment or Follow-up F3 Vizzutti et al. Gut 2009;58:156-60.
    15. 15. Shear wave propagation velocity according to severity of hepatic fibrosis (Metavir score) Sandrin L. Ultrasound Med Biol 2003 ; 29 : 1705 – 1713. E = 3.0 kPa F 0 E = 7.7 kPa F 2 E = 27 kPa F 4
    16. 16. Liver stiffness for each Metavir stage in CHC Box-and-whiskers plot Vertical axis is in logarithmic scale (wide range of F4 values) Gastroenterology 2005; 28:343 – 350.Hepatology 2005;41:48 – 54.
    17. 17. Correlation between LSM & fibrosis stage * Gastroentérol Clin Biol 2008;32,58-67. ** J Hepatol 2009;49:1062-68, Aliment Pharmacol Ther 2008;28:1188-98. *** Hepatology 2010;51:454-62. Gastroentérol Clin Biol 2008;32:58-67.
    18. 18. Accuracy of a diagnostic test • Dichotomous test (only 2 results) Sensibility (Sn) Specificity (Sp) Positive Predictive Value (PPV) Negative Predictive Value (NPV) Likelihood Ratios + & – (LRs) Diagnostic Odds Ratio (OR) • Multilevel test (> 2 results) Receiver Operating Characteristic (ROC) Newman TB & Kohn MA. Evidence-based diagnosis. Cambridge University Press, Cambridge, UK, 1st edition, 2009. CIs
    19. 19. Hypothetical ROC curve Pines JM & Everett WW. Evidence-Based emergency care: diagnostic testing & clinical decision rules. Blackwell’s publishing, West Sussex, UK, 2008.
    20. 20. Accuracy of diagnostic test using AUC of ROC Value Accuracy 0.90 - 1.00 Excellent Pines JM & Everett WW. Evidence-Based emergency care: diagnostic testing & clinical decision rules. Blackwell’s publishing – West Sussex – UK – 2008. AUROC of a ‘‘good” test should be ≥ 0.80 0.80 - 0.90 Good 0.70 - 0.80 Fair 0.60 - 0.70 Poor
    21. 21. Meta-analysis of TE for staging liver fibrosis Severe fibrosis (F3): 0.89 (95% CI: 0.88 – 0.91) Friedrich R et al. Gastroenterology 2008 ; 134 : 960 – 974. Cirrhosis (F4): 0.94 (95% CI: 0.93 – 0.95) Cut-off value: 13.0 kPa Significant fibrosis (F2): 0.84 (95% CI: 0.82 – 0.86) Cut-off value: 7.7 kPa 50 studies – random effect – all type of CLD
    22. 22. Significance of wide range of LSM in cirrhosis 13 - 75 kPa Ascites HCC ? Variceal bleeding Foucher J et al. Gut 2006 ; 55 : 403 – 408. EV stage 2 or 3 26 Child-Pugh B or C 36 49 53 622.5 7513
    23. 23. Cumulative incidence of HCC based on LSM 866 CHC – Mean follow-up 3 years LSM: Liver Stiffness Measurement – HR: Hazard Ratio Masuzaki R et al. Hepatology 2009 ; 49 : 1954 – 1961. LSM > 25 kPa HR 45.5 (p< 0.001) LSM ≤ 10 kPa HR 0 10 < LSM ≤ 15 kPa HR 16.7 (p< 0.001) 15 < LSM ≤ 20 kPa HR 20.9 (p< 0.001) 20 < LSM ≤ 25 HR 25.6 (p< 0.001)
    24. 24. Reproducibility of TE in assessing hepatic fibrosis. Bland Altman Plot Fraquelli M et al. Gut 2007 ; 56 : 968 – 973. 200 patients with chronic liver disease 2 different operators within 3 days (800 exams) 8 patients scored outside limits of agreement Upper limit Lower limit Mean 95% limit of agreement
    25. 25. Cost of FibroScan® versus liver biopsy • Liver biopsy* Cost of liver biopsy 703 – 1 566 € in a French hospital with a one day observation period • Fibroscan® ** FibroScan equipment 70 000 € Low running cost except probe calibration twice/year Cost per FibroScan exam 100 € with 150 exams annually * Blanc J et al. Hepatol Res 2005 ; 32 : 1 – 8. ** Canadian Agency for Drugs and Technologies in Health (CADTH). Transient Elastography (FibroScan) for Non-invasive Assessment of Liver Fibrosis; 2006.
    26. 26. Liver biopsy size • Because grading, & staging of nonneoplastic diffuse parenchymal liver disease is dependent on adequate sized biopsy, a biopsy of at least 2-3 cm in length & 16-gauge in caliber is recommended • Presence of fewer than 11 complete portal tracts in pathology report may be incorrect in recognition of grading, & staging due to insufficient sample size AASLD guidelines. Hepatology, 2009 ; 49 : 1017 – 1044.
    27. 27. Limitations of liver biopsy • Sampling errors Extremely small portion of liver (1/50 000) • Intraobserver & interobserver variation Even when widely validated systems used for score • Invasive procedure Morbidity: pain in 20% of patients Major complications: bleeding or hemobilia in 0.5% Mortality:
    28. 28. Grading & staging systems for chronic hepatitis IASL1 Batts–Ludwig2 Metavir3 1 Desmet VJ et all. Hepatology 1994;19:1513-1520. 2 Batts KP et all. Am J Surg Pathol 1995;19:1409-1417. 3 Bedossa P et all. Hepatology 1996;24:289-293. Grading system Minimal activity Mild activity Moderate activity Marked activity Marked activity & bridging Grade 1 Grade 2 Grade 3 Grade 4 Grade 4 A1 A2 A3 A3 A3 Staging system No fibrosis Fibrous portal expansion Few bridges or septa Numerous bridges Cirrhosis Stage 0 Stage 1 Stage 2 Stage 3 Stage 4 F0 F1 F2 F3 F4 (kappa 0.2 – 0.6) (kappa 0.5 – 0.9)
    29. 29. kappa score ≥ 0.6 indicates good agreement Interpretation of different values of kappa Kappa from Greek letter κ Value of kappa Strength of agreement 0 – 0.20 Poor 0.21– 0.40 Fair 0.41– 0.60 Moderate 0.61– 0.80 Good 0.81–1.00 Very good Perera R, Heneghan C & Badenoch D. Statistics toolkit. Blackwell Publishing & BMJ Books, Oxford, 1st edition, 2008.
    30. 30. Liver biopsy is not the “gold standard” but is the “best available gold standard”
    31. 31. Contraindications of liver biopsy • Uncooperated patients • Disorders in coagulation profile • Severe ascites • Cystic lesions • Vascular tumors (hemangiomas) • Amiloidosis • Congestive liver disease
    32. 32. R0C curves for FibroScan, FibroTest, & APRI for cirrhosis (F0 – F3 vs F4) Castera L et al. Gastroenterology 2005 ;128 : 343 – 50. Castera L et al. Lancet 2010 ; 375 : 1419 – 20.
    33. 33. Pitfalls of liver stiffness measurement  Obesity  Operator experience  Acute liver injury  Extrahepatic cholestasis  Increased CVP  Ascites  Narrow intercostal spaces
    34. 34. Obesity & operator experience
    35. 35. Limitations of liver stiffness measurement 13 369 examinations – 5 year prospective study – 5 operators BMI > 30 kg/m2 (OR 7.5) Operator experience (OR 2.5) Age > 52 years (OR 2.3) Type 2 diabetes (OR 1.6) Failure (3%) BMI > 30 kg/m2 (OR 3.3) Operator experience (OR 3.1) Age > 52 years (OR 1.8) Female sex (OR 1.4) Hypertension (OR 1.3) Type 2 diabetes (OR 1.1) Unreliable results (16%) Castéra L et al. Hepatology 2010 ; 51 : 828 – 835. LSM uninterpretable in one of five cases Main raisons: obesity ( WC) – operator experience
    36. 36. Failure rates according to BMI 7261 patients at the time of first examination Castéra L et al. Hepatology 2010 ; 51 : 828 – 835.
    37. 37. Unreliable results according to BMI 6968 patients at the time of first examination Castéra L et al. Hepatology 2010 ; 51 : 828 – 835.
    38. 38. Feasibility of LSM with FibroScan® using XL probe New probe for obese patients de Lédinghen V et al. Liver International 2010 ; : 1043 – 1048. 60% not measured by M probe successfully measured by XL probe
    39. 39.  Acute liver injury
    40. 40. Acute viral hepatitis increases liver stiffness 18 patients with acute viral hepatitis I Peak increase in aminotransferase II Aminotransferase ≤ 50% of the peak III aminotransferase levels ≤ 2 ULN Arena U et al. Hepatology 2008 ; 47 : 380 – 384.
    41. 41. Acute viral hepatitis increases liver stiffness 18 patients with acute viral hepatitis I Peak increase in aminotransferase II Aminotransferase ≤ 50% of the peak III aminotransferase levels ≤ 2 ULN Arena U et al. Hepatology 2008 ; 47 : 380 – 384.
    42. 42.  Extrahepatic cholestasis
    43. 43. Obstructive jaundice due to GIST occluding CBD Millonig G et al. Hepatology 2008 ; 48 : 1718 – 1723. Stent occlusion Bilirubin 8 mg/dL Stiffness 10 kPa Bilirubin 3.5 mg/dL Stiffness 5.7 kPa Stent placement Bilirubin 2 mg/dL Stiffness 5 kPa
    44. 44. Liver stiffness as a function of bile duct ligation 10 German landrace pigs: 5 controls – 5 BD ligation Millonig G et al. Hepatology 2008 ;48 : 1718 – 1723. 4.6 kPa 8.8 kPa 6.1 kPa
    45. 45.  Increased CVP
    46. 46. Representation of clamping site of the IVC 5 German landrace pigs Millonig G et al. J Hepatol 2010 ; 52 : 206 – 210. Experiment approved by local committee for Animal Welfare University of Heidelberg – Germany
    47. 47. LSM after clamping & reopening of IVC 5 anesthesized landrace pigs Millonig G et al. J Hepatol 2010 ; 52 : 206 – 210. P < 0.001 P < 0.001 Before clamping 3.1 kPa 5 min after reopening 5.1 kPa 27.8 kPa 5 min after clamping
    48. 48. Liver stiffness directly influenced by CVP 10 patients with CHF before & after recompensation Millonig G et al. J Hepatol 2010 ; 52 : 206 – 210. Median 40.7 Median 17.8 p = 0.004
    49. 49.  Ascites
    50. 50. Ascites in liver cirrhosis Ascites grade 1: detectable only by ultrasound
    51. 51.  Narrow intercostal spaces
    52. 52. Position of FibreScan® probe Dorsal decubitus position Right arm in maximal abduction
    53. 53. Interpretation of the results of LSM should always be done by expert clinicians according to clinical context
    54. 54. Transient elastography in clinical practice Examination quality 10 shots at least Success rate ≥ 60% IQR ≤ 25% of median value Liver disease Not used in acute hepatitis Not used in acute exacerbation Not used in ascites & EH cholestasis Choice of cutoff point Cutoffs different for each CLD Range of value rather than cutoff De Lédinghen V et al. Gastroentérol Clin Biol 2008 ; 32 : 58 – 67.
    55. 55. Questions
    56. 56. Thank You

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