Role of Ultrasound in Chronic Liver Disease: Is There Something New?

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1 Med. J. Cairo Univ., Vol. 85, No. 1, March: , Role of Ultrasound in Chronic Liver Disease: Is There Something New? WAFAA M. ABDALLAH, M.Sc.*; SHERIF F. ABD EL-RAHMAN, M.D.**; REDA S. ABD EL-LATIF, M.D.** and GAMAL ESMAT, M.D.*** The Department of Diagnostic & Interventional Radiology, National Hepatology & Tropical Medicine Research Institute* and Faculty of Medicine, Cairo University** and The Department of Endemic Medicine & Hepatology, Faculty of Medicine, Cairo University***, Egypt Abstract Background: Ultrasound is a non-invasive, widely used imaging method in the assessment and follow-up of patients with chronic liver disease. New advances in ultrasound such as Real Time Elastography (RTE) have been studied as a recent non-invasive method for liver fibrosis assessment. Aim of the Study: To determine the diagnostic accuracy of routine clinical ultrasound in the staging of liver fibrosis in chronic viral hepatitis, and assess the possible role of RTE as a new method of quantitative analysis of liver fibrosis. Methods: This study included sixty patients with Chronic Hepatitis C (CHC) and compensated cirrhosis that underwent liver biopsy. Conventional abdominal ultrasound and RTE were performed to all patients in addition to a group of twenty healthy volunteers. Findings were compared to the pathological liver fibrosis staging. Results: The sensitivities of three ultrasound features (surface nodularity, liver edge, and parenchymal echotexture), assembled as a grading system, were evaluated as an indicator of the presence of liver fibrosis. Sensitivities were: 63%, 22% and 29.4% for F1, F2 and F3/4 respectively. F3/4 group was further evaluated for another feature (caudate/right lobe ratio) and cirrhosis (F4) was diagnosed with a sensitivity 75%. Conventional ultrasound was able to reach a correct diagnosis in regard to histological fibrosis stage in 44.4% of cases. Meanwhile, RTE highly correlated with the stage of liver fibrosis (r=0.746, p<0.001), with a high diagnostic accuracy for F>3 (AUROC 0.96, sensitivity 94%, and specificity 93%), and for F >!2 (AUROC 0.93, sensitivity 84.6%, and specificity 85.4%), however, it showed a lower accuracy for F 1 (AUROC 0.87, sensitivity 83%, and specificity 71.4%). Conclusion: Conventional US cannot differentiate accurately the different fibrosis stages, yet, it can be useful in confirming presence of cirrhosis. RTE can be a new reliable tool for liver fibrosis assessment in patients with CHC, being able to differentiate absence and mild fibrosis from advanced fibrosis and cirrhosis. Correspondence to: Dr. Wafaa M. Abdallah, drwafaamahmoud@hotmail.com Key Words: Liver fibrosis Chronic hepatitis C Ultrasound Real time elastography. Introduction CHRONIC hepatitis C is the most common cause of liver fibrosis. Hepatitis C Virus (HCV) infects approximately 170 million individuals worldwide, the prognosis and management of these patients depend on the degree of liver fibrosis. Therefore, the assessment of liver fibrosis is an important determinant in these patients. Because of the known disadvantages of Liver Biopsy (LB) e.g. invasive and under the fact that absolute staging of hepatic fibrosis has become less important with the new trend to treat all CHC patients, owing to the new drug regimens, the international guidelines recommended that non-invasive methods for hepatic fibrosis assessment can be used in routine clinical practice [1-3]. Ultrasound is an easy, non-invasive imaging method that is widely available and routinely used in evaluation of patients with chronic liver disease. Previous studies have shown that routine clinical US has good sensitivity and high specificity for detecting cirrhosis, and a limited ability in detecting significant fibrosis [4]. Image characteristics of hepatic fibrosis/cirrhosis includes; surface nodularity, heterogeneous echo texture, decrease in the volume of right lobe, increase in the volume of caudate and left lobe, and narrowing of the hepatic veins [5]. However, several factors can affect the diagnostic accuracy such as inter- and intraobserver variability, technical level of the operator, interference of obesity, ascites and intestinal gas, modulation of the apparatus, and the complex pattern of changes in chronic liver disease that includes 393

2 394 Role of Ultrasound in Chronic Liver Disease: Is There Something New? mixed features of steatosis, necrosis, and inflammation [6]. Variable attempts were made to increase the sensitivity of ultrasound in staging of liver fibrosis. Several investigations of the utility of Doppler for assessing hepatic fibrosis have been made. However, correlation between Doppler indices and disease stage was uncertain, with no significant differences observed with increasing severity of liver disease [7]. Contrast-enhanced ultrasound and Hepatic Vein Transit Time (HVTT) in which, reduced HVTT suggested the presence of intrahepatic shunting that is found with advanced hepatic fibrosis [8]. US tissue characterization has also been attempted in the literature for objective evaluation of tissue by analyzing the signals obtained from the US system e.g. intensity of radiofrequency signals, and extent of scattering [5]. In this context, liver elastography is a new proposed method for assessment of liver fibrosis. The rational for liver elasticity measurement was based on the simple basic concept that the hardness of the liver tissue is related to the degree of liver fibrosis as noted on palpation [9]. Ultrasound elastography has two known types: Shear-wave elastography and real-time (strain) elastography. Shearwave based techniques including transient elastography measure the propagation of shear wave induced within the tissues to estimate liver stiffness [10], however, RTE is technically different as it compares and analyses echo signals before and after compression [9], then the relative strain of tissues is displayed as a colored strain image (elastogram) overlying the B-mode image [10]. Strain images show a progressively increasing patchy pattern as fibrosis progresses [11], however, to increase objectivity, different methods of elastogram analysis have been developed to retrieve a semi-quantitative estimate of liver fibrosis [12]. The usefulness of RTE for assessing liver fibrosis is well established in literature [11,13], however, variable results have been reported about the performance of the different quantitative methods [6], which limits the recommendation for its use in clinical practice [14]. So since ultrasound is a routine clinical investigation for patients with chronic liver disease, we sought to determine its diagnostic accuracy and the reliability of the new additional RTE mode in predicting the stages of liver fibrosis in CHC patients through comparing the results to LB as the reference standard. Patients and Methods Sixty patients diagnosed with CHC or cirrhosis and underwent liver biopsy at the National Hepatology and Tropical Medicine Research Institute, during the year of 2014, were included in the study. Patients with established cirrhosis were not scheduled for liver biopsy. We excluded cases with coinfection, decompensated liver cirrhosis or hepatic focal lesions. Twenty healthy volunteers were also included in the study as a control group. No liver biopsy was performed and they were considered as Metavir F0 stage. Liver histology: US-guided LB was performed using a semiautomatic true-cut needle (16G). The specimen lengths ranged from 10 to 25mm and the sections were stained with hematoxylin and eosin, and Masson trichrome staining. We used the METAVIR scoring system for liver fibrosis staging and activity grading. Fibrosis: F0=no fibrosis; F 1=portal fibrosis without septa; F2=portal fibrosis with few septa; F3=numerous septa without cirrhosis; and F4= cirrhosis, and inflammatory activity: A0=no histologic activity; A1=mild activity; A2=moderate activity; and A3=severe activity. Ultrasound examination: Examinations were performed using HI VISION Preirus (Hitachi Medical Corp., Japan) that includes a convex probe (3.5-5MHz) and a superficial probe (7-12MHz) for evaluation of the liver surface. Real time tissue elastography: RTE was performed using HI VISION Preirus (Hitachi Medical Corp., Japan) with a EUP-L52 linear probe (7-3MHz). Internal compression and relaxation induced by heart beat was used to obtain the strain images. Liver was scanned from the right intercostal space to measure tissue elasticity within the right hepatic lobe. The ROI (2.5 X 2.5cm) for RTE was set at a depth of more than 1 cm from the surface of the liver. Analysis of RTE image features were automatically performed with computer software. Multiple regression analysis was then performed to calculate the liver fibrosis index using the following equation (Table 1): LFI = X MEAN X SD X %AREA X COMP X SKEW 0.28x KURT X ENT X IDM X ASM 5.54 Statistical analysis: Data were statistically described in terms of mean ± standard deviation, median and range, or frequencies and percentages. Comparison of nu-

3 Wafaa M. Abdallah, et al. 395 merical variables between the study groups was done using One Way ANOVA and Kruskal Wallis test. Correlation between various variables was done using Spearman rank correlation equation. Accuracy was represented using the terms sensitivity, and specificity. Receiver Operator Characteristic (ROC) analysis was used to determine the optimum cut off value for the studied diagnostic markers. p-values less than 0.05 was considered statistically significant. All statistical calculations were done using computer program SPSS (Statistical Package for the Social Science; SPSS Inc., Chicago, IL, USA) release 15 for Microsoft Windows (2006). Results The study included 80 subjects; however, 6 patients were excluded from our study because of unreliable RTE images (unable to hold breath "bronchial asthma", weak cardiac pulsations and multiple reflection artifacts). The indicated stages of hepatic fibrosis in the study group were: F0 in 21 subjects, F1 in 27 subjects, F2 in 9 subjects, F3 in 13 subjects and F4 in 4 subjects. Steatosis was noted in 33 patient (66%) of patients underwent LB. Analysis of routine clinical hepatic ultrasound: For the study purpose, three features (surface nodularity, liver edge, and parenchymal echotexture) were evaluated. We assessed the sensitivity of these features with a grading system (Table 2) modified from a previously published scoring system reported by Nishiura et al., [15] and Choong et al., [16] groups. A normal liver with no fibrosis was expected to receive a grade 0 for all three features. A liver with mild fibrosis (Grade 1) would be expected to have either mild surface nodularity &/or altered echotexture ± mild blunting of the liver edge. Moderate liver fibrosis (Grade 2) was expected to show some surface nodularity and mild/moderate coarsening of echotexture ± blunting of the liver edge. In severe fibrosis (Grade 3), the liver was expected to appear more nodular and show coarse echotexture changes with blunting of liver edge. Finally, cirrhotic livers were expected to demonstrate all the above features in addition to other features as small right lobe with left and caudate lobe hypertrophy ± signs of decompensation. Correlation of ultrasound grades with staging of fibrosis: When considering US grade 0 as a normal liver with no fibrosis corresponding to F0, Grade 1 as a mild liver fibrosis corresponding to F1, Grade 2 as a moderate liver fibrosis corresponding to F2 (significant fibrosis) and Grade 3 as sever fibrosis corresponding to F3/4 (advanced fibrosis and cirrhosis), there were; (Tables 3,4) (17/27) F1 cases (=63%) diagnosed as Grade 1, (10/27) (=37%) were underestimated as Grade 0, no overestimated cases. (2/9) F2 cases (=22%) diagnosed as Grade 2, (7/9) (=78%) were underestimated as Grade 1 (6 cases) and Grade 0 (1 case), no overestimated cases. (5/17) F3/4 cases (=29.4%) diagnosed as Grade 3, (12/17) (=70.6%) were underestimated as Grade 2 (8 cases) and Grade 1 (4 cases), no overestimated cases. This group was further evaluated for another feature (caudate/right lobe ratio) and liver cirrhosis was diagnosed in 3 cases. Thus (3/4) F4 cases (=75%) were diagnosed as cirrhosis and 1 case was underestimated as Grade 2. So conventional ultrasound was able to reach a correct diagnosis in regard to histological fibrosis stage in 24/54 cases (=44.4%). Correlation of RTE with staging of fibrosis: The Spearman's correlation coefficient showed a high correlation between the LFI and the stage of liver fibrosis (r=0.746, p<0.00 1). The median LFI in patients with F0, F1, F2, F3, and F4 were 1.6, 2.1, 2.46, and 4, respectively. LFI showed a stepwise increase with increasing the histologic severity of fibrosis. F1 did not show a statistically significant difference in LFI from F2 stage, the same was observed for F3 and F4 stages, whereas for all other combinations of stages, LFI significantly differed with the stage of liver fibrosis ( p <0.001). The overall AUROC for F0 versus F1-4, F0-1 versus F2-4, and F0-2 versus F3-4 were 0.876, and respectively. When the cutoff value of LFI was set to 1.65 for F0 versus F1-4, the sensitivity, specificity and accuracy were 83, 71.4 and 79.7%, respectively. The PPV and NPV were 88 and 62.5%, respectively. For F0-1 versus F2-4 when the cutoff value was 2.4, the sensitivity, specificity and accuracy were 84.6, 85.4 and 85%,

4 396 Role of Ultrasound in Chronic Liver Disease: Is There Something New? respectively. The NPV to exclude F2 or greater stage was 91% and the PPV was 75.9%. For F0-2 versus F3-4, when the cutoff value was 2.96, the sensitivity, specificity and accuracy were 94, 93 and 93%, respectively. The NPV to exclude advanced fibrosis was 98% and the PPV was 80% (Table 5). The Spearman's correlation coefficient showed no significant correlation between LFI and the degree of necro-inflammation ( r=0.419, p=0.002). Hepatic steatosis was classified into 3 groups as no steatosis (n=17), 0%-30% (n=24), and >30% (n=9). The Spearman's coefficient showed no significant correlation between LFI and steatosis ( r= 0.385, p=0.006). Table (1): Feature values [5]. MEAN SD %AREA Mean of relative strain value within the ROI Standard deviation of relative strain value within the ROI Area of low strain (blue) within the ROI Table (3): Correlation of ultrasound features and histological grading of liver. Ultrasound feature Surface nodularity: Histological grade of fibrosis METAVIR F1 F2 F3 F4 Smooth Mildly irregular 4 1 Irregular 1 1 Highly irregular 1 Edge: Sharp 11 1 Mildly blunted Blunted Echotexture: Normal Mildly coarse Moderately coarse Highly coarse 1 3 COMP SKEW KURT Complexity of low strain (blue) area within the ROI =(perimeter)2/area Skewness - asymmetry of the histogram Kurtosis - peakedness of the histogram Grade: ENT IDM ASM Entropy - textural complexity Inverse difference moment - textual local homogeneity Angular second moment - textual homogeneity Table (2): US grades of liver fibrosis. Grade 0: Grade 1: Grade 2: Grade 3: Surface: Smooth Liver edge: Sharp Parenchymal echotexture: Normal Surface: Mildly irregular (superficial probe) Liver edge: Mildly blunted Parenchymal echotexture: Mildly coarse Surface: Irregular Liver edge: Blunted Parenchymal echotexture: Moderately coarse Surface: Highly irregular Liver edge: Blunted Parenchymal echotexture: Highly coarse Table (4): Correlation of ultrasound grades and histological grading of liver. Stage Grade F 1 (27) 10 (37%) 17 (63%) F2 (9) 1 (11.1%) 6 (66.7%) 2 (22.2%) F3 (13) 4 (30.8%) 7 (53.8%) 2 (15.4%) F4 (4) 1 (25.0%) 3 (75.0%) Table (5): Cutoff values of LFI for the determination of fibrosis stage. F1 F2 F3 Cut-off AUC Sensitivity (%) Specificity (%) Accuracy (%) PPV NPV

5 Wafaa M. Abdallah, et al. 397 Region to analyze for Strain Histogram measurement Strain graph Fig. (1): Normal liver. The color-coded elastogram shows a diffuse soft pattern, characterized by a homogeneous light-green image. Fig. (2): Display data list to check the measurement result of 11 types of feature amount. Fig. (3): Example of liver RTE image and result of LFI (F2 stage): The color-coded elastogram shows an intermediate pattern, characterized by a mottled image with blue spots on a light-green background. LFI: Fig. (4): Example of liver RTE image and result of LFI (F3 stage): The color-coded elastogram shows the hard pattern, characterized by image patchiness of green, and blue. LFI: Discussion In this study we tried to evaluate the diagnostic accuracy of routine US in the staging of liver fibrosis in CHC patients, we specifically assessed three features (surface nodularity, liver edge, and parenchymal echotexture). Our results showed that conventional US had an overall accuracy of 44.4% in staging liver fibrosis; however, the presence of either feature is highly specific in predicting significant fibrosis. In our study all wrongly diagnosed cases were due to underestimation of the fibrosis stage, implicating that the patient can have significant, advanced or even cirrhosis and still remain unrecognized which can affect his management plane and post treatment surveillance. So we believe that conventional US cannot differentiate accurately the different fibrosis stages and it can be useful in confirming presence of cirrhosis or for stage F4 patients who are usually on surveillance for complications, such as HCC and portal hypertension. Yet a limitation of our study is that the control group (F0) were not enrolled among patients at time of initial evaluation and they were assessed as an individual group so the ability of conventional US to rule out or detect early fibrosis was not assessed, this might also explain higher accuracy in detecting F 1 stage and might also slightly increased the overall accuracy. We also assessed a newly developed ultrasound elastography modality that reflects liver elasticity in the form of Liver Fibrosis Index (LFI), which has been evaluated in multiple previous studies Tatsumi et al. [17], Ferraioli et al. [18], Fujimoto et al. [5], Tomeno et al. [19], Yada et al. [20], Tamaki et al. [21] and Kim et al. [22]. In our study LFI highly correlated with the stage of liver fibrosis (r=0.746, p<0.001), with a high diagnostic accuracy for F >_3 (AUC 0.96, sensitivity 94%, specificity 93%), excluding advanced fibrosis with an NPV of 98%, and for F >_2 (AUC 0.93, sensitivity 84.6%,

6 398 Role of Ultrasound in Chronic Liver Disease: Is There Something New? specificity 85.4%), meanwhile it showed a lower accuracy for F >_ 1 (AUC 0.87, sensitivity 83%, specificity 71.4%). A partial overlapping of LFI values was observed, particularly for (F1 and F2) and for (F3 and F4) stages, indicating a lower efficiency in discriminating between the contiguous stages of hepatic fibrosis. Advanced stage of chronic liver disease includes cases previously diagnosed as cirrhosis (METAVIR F4), and shortly falls backwards to include METAVIR F3 cases with architectural distortion [23]. Both are known to have the worst survival curves to a similar extent [24]. On the basis of these data, we considered stage 3 and 4 fibrosis as a single class in our study. This has enabled to increase the accuracy of RTE to identify cirrhosis and has decreased the false-negative rate in the present series, which allowed for identification of all patients with the worst prognosis. RTE was successfully performed in overweight and obese patients. Our study did not show any association between BMI and LFI measurements, neither results were influenced by inflammatory activity nor hepatic steatosis. So, RTE can be performed in cases for which the results of Transient elastography (fibroscan) are considered unreliable. And, combined with serum markers, RTE can be used as an adjunct US tool in the initial assessment of liver fibrosis to exclude the presence of significant or advanced liver fibrosis. Results are instantaneous and the clinician can use them to make decisions during patients' visits. When both tests indicate mild or no disease, then the combined result should be both sensitive and specific, and clinicians can be confident in this result. Likewise, when both tests indicate advanced or significant disease, this result has high sensitivity, high specificity, and a high predictive value. Still, RTE can have limitations related to being operator dependent. Training is needed to avoid artifacts related to obesity, to set the ROI not to include vessels or rib shadowing and to adjust the position of the probe to image the liver where compression/relaxation is homogeneous and axial to the probe. Study limitations: The distribution of patients in our study was not equal through METAVIR scores, 36.5% of subjects were in the F1 stage, but since the studied population was potential candidates to antiviral therapy, we think that the uneven distribution of patients reflects what is seen in clinical practice. Conclusion: Ultrasound is a widely used modality, easy to use and cost-effective, however it is not reliable for staging of liver fibrosis and for making therapeutic decisions in cases of CHC patients. But it is still of great value in follow-up and surveillance of patients with advanced fibrosis. The new application of US elastography increases sensitivity and can be used for diagnosing liver fibrosis in patients with CHC. RTE showed a high performance and proved to be especially valuable for diagnosis of advanced hepatitis and early cirrhosis that are concerns in clinical practice. Future studies on larger patient cohorts will be necessary for the validation of RTE, and to study whether or not RTE has the potential to substitute for TE. Conflict of interest: None declared. References 1- European Association for the Study of the Liver. EASL Clinical Practice Guidelines: Management of hepatitis C virus infection. J. Hepatol., Aug., 55 (2): , MYERS R.P., RAMJI A., BILODEAU M., WONG S. and FELD J.J.: An update on the management of hepatitis C: Consensus guidelines from the Canadian Association for the Study of the Liver. Can. J. Gastroenterol., Jun., 26 (6): , SHIHA G., SARIN S.K., IBRAHIM A., OMATA M., KUMAR A., LESMANA L.A., et al.: Liver fibrosis: Consensus recommendations of the Asian Pacific Association for the Study of the Liver (APASL). Hepatol. 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8 400 Role of Ultrasound in Chronic Liver Disease: Is There Something New?