In China, chronic hepatitis B (CHB) is the most common cause of liver fibrosis. Early and timely diagnosis of liver fibrosis and assessment of its progress can guide clinical decision-making, which is conducive to improving patient prognosis and improving patient quality of life. Liver biopsy (LB) is currently the "gold standard" for assessing the degree of liver fibrosis in the clinic, but it is an invasive procedure with unavoidable risks such as bleeding, sampling errors, and interpretation errors [1,2,3]. In recent years, non-invasive testing methods used clinically to assess the degree of liver fibrosis have received increasing attention, among which transient elastography (TE) technology is gradually being popularized, and FibroTouch and FibroScan are the two most used examination instruments. Previous studies have shown that TE is more effective in diagnosing the degree of liver fibrosis, but elevated levels of alanine aminotransferase (ALT) may lead to an increase in liver stiffness measurements (LSM), which in turn affects TE test results and the reliability of TE diagnostic liver fibrosis [4,5]. This study used FibroTouch to measure LSM in CHB patients and group them according to ALT levels, aiming to clarify the effect of ALT levels on FibroTouch's detection of LSM in CHB patients.
The research is innovative:
In view of the fact that there are few studies to explore the effect of varying degrees of elevated alanine aminotransferase (ALT) levels, especially mildly elevated ALT levels, on the degree of liver fibrosis in patients with chronic hepatitis B (CHB), this study grouped 145 patients with CHB according to the ALT level, and divided the liver puncture biopsy results into mild liver fibrosis (F1), significant liver fibrosis (F2-F3), and cirrhosis (F4) according to the METVIR scoring system standards. Finally, by comparing the liver hardness value (LSM) measured by FibroTouch in patients with CHB with different ALT levels and different degrees of liver fibrosis, and plotting the subject work characteristic (ROC) curve, it was found that fibroTouch detection of LSM in CHB patients with ALT <2 × reference value (ULN) was not significantly affected, while FibroTouch detection of CHB patients with LSM at ALT ≥2× ULN may be overestimated. This is of certain reference value for clinicians to analyze the LSM detected by FibroTouch in CHB patients and to judge the degree of liver fibrosis in combination with the ALT level.
1 Objects and methods
1.1 Research Subjects
From May 2016 to March 2019, 145 CHB patients were selected from the Department of Infectious Diseases ward of the Second Affiliated Hospital of Dalian Medical University, all of which met the CHB diagnostic criteria in the Guidelines for the Prevention and Treatment of Chronic Hepatitis B (2019 Edition)[6] jointly formulated by the Infectious Diseases Branch of the Chinese Medical Association and the Hepatology Branch of the Chinese Medical Association in 2019, of which 89 were males and 56 were females, and the median age was 44 (35,55) years old. Exclusion criteria: (1) ALT≥5× upper limit of normal (ULN), decompensated stage of cirrhosis, primary liver cancer; (2) hepatitis A, hepatitis C, hepatitis E, autoimmune liver disease, drug-induced hepatitis, fatty liver disease, AIDS, bile duct obstruction; (3) alcoholism or body mass index ≥28 kg/m2; (4) there is a sign of contraindication to liver puncture such as bleeding tendencies or disagreement with LB; (5) other systemic serious diseases ;(6) Myocardial injury; (7) History of malignant tumors; (8) Maternal conditions; (9) Recent history of right upper quadrant trauma and unhealed wounds; (10) Lack of capacity for civil conduct or limited capacity for civil conduct. This study is a retrospective study, which was reviewed and approved by the Second Affiliated Hospital of Dalian Medical University, and all patients signed the informed consent form and performed LB and completed serology and FibroTouch examination within 1 week.
1.2 Grouping and staging
According to the ALT level, all patients were divided into 46 cases in group A (ALT<1×ULN), 64 cases in group B (1×ULN≤ ALT < 2 ×ULN), and 35 cases in group C (2 × ULN≤ ALT<5 ×ULN). According to the METAVIR scoring system standard, the LB results were divided into mild liver fibrosis (F1), significant liver fibrosis (F2-F3), cirrhosis (F4), 16 patients in group A were F1, 22 patients were F2-F3, and 8 patients were F4; 14 patients in group B were F1, 44 patients were F2-F3, 6 cases were F4, 6 patients in group C were F1, 22 patients were F2-F3, and 7 patients were F4. The grouping and staging of the study subjects are detailed in Figure 1.
Figure 1 Grouping and staging of 145 CHB patients
Figure 1 Grouping and staging of the research objects
1.3 Serological tests
All patients were sampled with 2 ml of fasting venous blood in the morning, and the ALT and total bilirubin levels were detected by Siemens automatic biochemical analyzer, of which the ALT reference value was≤ 40 U/L for men and 35 U/L for ≤ for women.
1.4 FibroTouch check
All patients are tested with FibroTouch produced by Wuxi Haiskell Medical Technology Co., Ltd., and the patient is taken in a flat lying position and the right arm is extended to the maximum extent; after the probe is applied with couplant, the patient's right axillary front line is selected to the 7th, 8th and 9th intercostal parts of the axillary line and close to the middle of the intercostal space, and the ideal position is selected to avoid the blood vessels under the ultrasound assisted by the instrument; after 10 consecutive successful tests, the median is taken as the final detection value, in kPa. The FibroTouch inspection requires a detection success rate of >60% and a variance of < 1/3 of the median; the operators are trained in the system.
1.5 LB
2 liver biopsies were taken from different parts of the liver of all patients with a 16G biopsy needle, each length ≥ 1.5 cm (≥ 8 complete sink areas), fixed with 10% formalin solution, paraffin-embedded, and stained with hematoxylum-eosin (HE). The film was read by two experienced senior pathologists alone, and the LB interpretation results were inconsistent after consultation and discussion.
1.6 Statistical methods
SPSS 20.0 statistics software was used for data analysis. If the measurement data is normally distributed, it is expressed as (x±s), using one-way ANOVA, and if it is biased, it is expressed as M(P25, P75), using a rank sum test. The counting data is expressed as a relative number and is tested by χ2. The correlation between overall LSM and the degree of liver fibrosis in three groups of patients was analyzed using a Spearman rank correlation analysis. Taking the LB results as the "gold standard", the subject work characteristic (ROC) curve was plotted and the area under the ROC curve (AUC) was calculated, and the diagnostic efficacy of FibroTouch on the degree of liver fibrosis in patients with CHB was analyzed, and the closer the AUC was to 1.0, the higher the diagnostic efficacy [7]. The difference in P<0.05 was statistically significant.
2 Results 2.1 Male proportion, age, total bilirubin level and LSM of patients with CHB levels with different ALT levels were compared
There was no significant difference between the male proportion and total bilirubin levels in the three groups (P>0.05), and the age, overall LSM and LSM of patients F1, F2-F3 and F4 in the three groups were statistically significant (P<0.05). The overall LSM and F1, F2-F3 and F4 patients in Group A and Group B were lower than those in Group C, and the difference was statistically significant (P<0.05). See Table 1 for details.
Table 1 Male proportion, age, total bilirubin level and LSM of patients with CHB with different ALT levels
Table 1 Comparison of male proportion，mean age，total bilirubin and liver stiffness measured by FibroTouchinchronic hepatitis B patients by ALT level
2.2 Male proportion, age, total bilirubin level and LSM of chB patients with different degrees of liver fibrosis
2.2.1 A组
There was no significant difference in male proportion, age and total bilirubin levels in group A (P>0.05) in F1, F2-F3 and F4, and there was a statistically significant difference in LSM in patients with F1, F2-F3 and F4 in group A (P<0.05). The differences were statistically significant in group A group in which F1 patients had LSM was lower than that of F2-F3 and F4 patients, and in F2-F3 patients with LSM was lower than that of F4 patients (P<0.05). See Table 2 for details.
Table 2 Male proportions, age, total bilirubin levels and LSM of patients with CHB with different degrees of liver fibrosis in group A
Table 2 Comparison of male proportion，mean age，total bilirubin and liver stiffness measured by FibroTouch in chronic hepatitis B patients in group A by liver fibrosis assessed by liver biopsy
2.2.2 Group B
There was no statistically significant difference in the male proportion, age and total bilirubin levels of patients F1, F2-F3 and F4 in Group B (P>0.05), and the difference was statistically significant (P<0.05) in LSM in Group B.B. The difference was statistically significant in group B group F1 patients with LSM lower than F2-F3 and F4 patients, and F2-F3 patients with F2-F3 patients with LSM lower than F4 patients (P<0.05). See Table 3 for details.
Table 3 Male proportion, age, total bilirubin level and LSM of patients with CHB with different degrees of liver fibrosis in Group B
Table 3 Comparison of male proportion，mean age，total bilirubin and liver stiffness measured by FibroTouch in chronic hepatitis B patients in group B by liver fibrosis assessed by liver biopsy
2.2.3 Group C
The male proportion, age and total bilirubin levels of patients with F1, F2-F3 and F4 in group C were not statistically significant (P>0.05); the difference was statistically significant in the LSM of patients F1, F2-F3 and F4 in group C (P<0.05). The differences were statistically significant in group C in F1 patients with LSM lower than F2-F3 and F4 patients, and F2-F3 patients with F2~F3 patients with LSM lower than F4 patients (P<0.05). See Table 4 for details.
Table 4 Comparison of male proportions, age, total bilirubin levels and LSM of CHB patients with different degrees of liver fibrosis in Group 4C
Table 4 Comparison of male proportion，mean age，total bilirubin and liver stiffness measured by FibroTouch in chronic hepatitis B patients in group C by liver fibrosis assessed by liver biopsy
2.3 Correlation Analysis
Spearman rank-related analysis showed that the overall LSM of the three groups was positively correlated with the degree of liver fibrosis (rs values were 0.81, 0.71, 0.73, P<0.001, respectively), as shown in Figure 2.
Figure 2 Correlation between LSM and degree of liver fibrosis in three groups of CHB patients
Figure 2 Correlations between LSM and liver fibrosis staging in the three groups of CHB patients
2.4 Diagnostic efficacy analysis
Drawing the ROC curve showed that the AUC of F2 to F3 in fibroTouch diagnosis population, group A, group B and group C was 0.904, 0.933, 0.914 and 0.897, respectively, and the AUC of F4 in the diagnosis of overall, group A, group B and group C was 0.942, 0.954, 0.989, 0.949 (P<0.001), respectively, as shown in Table 5, Figure 3, and Figure 4.
Fig. 3 ROC curve of FibroTouch diagnosis of F2 to F3 in patients with CHB
Figure 3 ROC curve for FibroTouch in diagnosing F2~F3 in patients with CHB
Figure 4 ROC curve of FibroTouch in diagnosing F4 in patients with CHB
Figure 4 ROC curve for FibroTouch in diagnosing F4 of patients with CHB
Table 5 Efficacy of FibroTouch in diagnosing the degree of liver fibrosis in patients with CHB
Table 5 Efficacy of FibroTouch in measuring liver fibrosis in patients with chronic hepatitis Bexplored using ROC analysis
3 Discussion
According to statistics, the number of chronic hepatitis B virus (HBV) infection in China is about 70 million [6]. When pathological evidence of significant hepatic fibrosis is clinically found in patients with CHB, antiviral therapy should be performed as soon as possible [8]. LB results are the "gold standard" for the diagnosis of liver fibrosis, but their clinical use is limited due to its invasiveness. Recent studies have shown that TE is the most effective and promising non-invasive method for clinically assessing the degree of liver fibrosis [9,10,11], but some studies have found that elevated ALT levels may affect the accuracy of LSM testing and the reliability of TE in diagnosing liver fibrosis [4,5]. Studies have shown that inflammation leads to increased liver stiffness, which in turn leads tees to overestimate liver fibrosis [5]. Possible mechanisms for increasing liver stiffness due to inflammation include tissue swelling, inflammatory infiltrates, edema, and increased intracellular pressure [12], but it is not clear how inflammation of varying severity affects LSM and the degree of liver fibrosis.
ALT is the most commonly used and direct indicator of liver inflammation in the clinic. In this study, CHB patients were divided into group A (ALT<1×ULN), group B (1×ULN≤ ALT<2×ULN), group C (2×ULN≤ ALT<5×ULN) according to the ALT level, and found that the overall LSM and F1, F2-F3 and F4 patients in group A and group B were lower than those in group C, and the possible reasons for the analysis were: significantly increased ALT levels (≥2×ULN) suggested that severe inflammation occurred in the liver. Severe inflammation can lead to an increase in liver hardness.
By further analyzing the LSM of CHB patients with different degrees of liver fibrosis, this study found that the LSM of F1 patients in group A, group B and group C was lower than that of F2-F3 and F4 patients, respectively, and the LSM of F2-F3 patients was lower than that of F4 patients, respectively; Spearman rank-related analysis results showed that the overall LSM of the three groups of patients was positively correlated with the degree of liver fibrosis, that is, the higher the degree of liver fibrosis, the higher the LSM of CHB patients detected by FibroTouch. Suggests that FibroTouch has a high diagnostic value for the degree of liver fibrosis in patients with CHB.
Liu Fang et al. [13] and Dai Wen et al. [14] found that fibroTouch's AUC for diagnosing significant liver fibrosis (≥ F2) in patients with CHB was 0.857 and 0.815, respectively, and the AUC for diagnosing cirrhosis (F4) was 0.875 and 0.920, respectively, suggesting that FibroTouch's diagnostic efficacy in cirrhosis of patients with CHB was better than that of liver fibrosis. In this study, it was found that the AUC of F2-F3 and F4 in the overall patients diagnosed by FibroTouch was 0.904 and 0.942, respectively, indicating that the heavier the degree of liver fibrosis in CHB patients, the higher the diagnostic efficiency of FibroTouch, which was consistent with the above research results. It should be pointed out that the AUC of FibroTouch in the diagnosis of group A and group B F2 to F3 was 0.933 and 0.914, respectively, which was higher than the AUC (0.897) of F2 to F3 in patients diagnosed in group C, suggesting that the ALT <2 × ULLN did not significantly affect the diagnostic efficacy of FibroTouch on the obvious liver fibrosis in CHB patients, and the ALT level was significantly increased (≥2× ULN) may result in a decrease in fibroTouch's diagnostic efficacy in patients with CHB for significant liver fibrosis, consistent with previous studies [4,5]; FibroTouch's AUC of F4 in patients with groups A, B, and C was 0.954, 0.989, and 0.949, respectively, suggesting that changes in ALT levels did not significantly affect FibroTouch's diagnostic efficacy of cirrhosis in patients with CHB, and were consistent with previous studies[15]. It was further confirmed that FibroTouch was better at diagnosing cirrhosis in patients with CHB than in liver fibrosis. Therefore, when clinically applying FibroTouch to detect LSM in CHB patients, we must pay attention to the increase in ALT levels and the actual detection value, and must be comprehensively judged to avoid overestimating the degree of liver fibrosis in CHB patients.
In summary, fibroTouch detection of LSM in CHB patients with ALT<2 × ULN was not significantly affected, while FibroTouch detection of CHB patients with LSM at ALT ≥2 × ULN may be overestimated and may not necessarily reflect the true degree of liver fibrosis, but the proportion of 2× ULN≤ ALT<5× ULN and F4 patients in this study is low and there are still differences in age between the three groups of patients, there is a certain selection bias and information bias, the results, The conclusions still need to be further verified in the future such as expanding the sample size and balancing the distribution of cases.
conflict of interest
There is no conflict of interest in this article.
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