Collagen co-localized with macrovesicular steatosis better differentiates fibrosis progression in non-alcoholic fatty liver disease mouse models.

Background: Non-alcoholic fatty liver disease (NAFLD) is a global commonly occurring liver disease. However, its exact pathogenesis is not fully understood. The purpose of this study was to quantitatively evaluate the progression of steatosis and fibrosis by examining their distribution, morphology, and co-localization in NAFLD animal models. Methods: Six mouse NAFLD groups were established: (1) western diet (WD) group; (2) WD with fructose in drinking water (WDF) group; (3) WDF + carbon tetrachloride (CCl4) group, WDF plus intraperitoneal injection of CCl4; (4) high-fat diet (HFD) group, (5) HFD with fructose (HFDF) group; and (6) HFDF + CCl4 group, HFDF plus intraperitoneal injection of CCl4. Liver tissue specimens from NAFLD model mice were collected at different time points. All the tissues were serially sectioned for histological staining and second-harmonic generation (SHG)/two-photon excitation fluorescence imaging (TPEF) imaging. The progression of steatosis and fibrosis was analyzed using SHG/TPEF quantitative parameters with respect to the non-alcoholic steatohepatitis Clinical Research Network scoring system. Results: qSteatosis showed a good correlation with steatosis grade (R: 0.823-0.953, p < 0.05) and demonstrated high performance (area under the curve [AUC]: 0.617-1) in six mouse models. Based on their high correlation with histological scoring, qFibrosis containing four shared parameters (#LongStrPS, #ThinStrPS, #ThinStrPSAgg, and #LongStrPSDis) were selected to create a linear model that could accurately identify differences among fibrosis stages (AUC: 0.725-1). qFibrosis co-localized with macrosteatosis generally correlated better with histological scoring and had a higher AUC in six animal models (AUC: 0.846-1). Conclusion: Quantitative assessment using SHG/TPEF technology can be used to monitor different types of steatosis and fibrosis progression in NAFLD models. The collagen co-localized with macrosteatosis could better differentiate fibrosis progression and might aid in developing a more reliable and translatable fibrosis evaluation tool for animal models of NAFLD.

The burden of hepatitis C virus in the world, China, India, and the United States from 1990 to 2019.

Background and aim: Hepatitis C virus infection can lead to an enormous health burden worldwide. Investigating the changes in HCV-related burden between different countries could provide inferences for disease management. Hence, we aim to explore the temporal tendency of the disease burden associated with HCV infection in China, India, the United States, and the world. Methods: Detailed data on the total burden of disease related to HCV infection were collected from the Global Burden of Disease (GBD) 2019 database. Joinpoint regression models were used to simulate the optimal joinpoints of annual percent changes (APCs). Further analysis of the age composition of each index over time and the relationship between ASRs and the socio-demographic Index (SDI) were explored. Finally, three factors (population growth, population aging, and age-specific changes) were deconstructed for the changes in the number of incidences, deaths, and DALYs. Results: It was estimated that 6.2 million new HCV infections, 0.54 million HCV-related deaths, and 15.3 million DALYs worldwide in 2019, with an increase of 25.4, 59.1, and 43.6%, respectively, from 1990, are mainly due to population growth and aging. China experienced a sharp drop in age-standardized rates in 2019, the United States showed an upward trend, and India exhibited a fluctuating tendency in the burden of disease. The incidence was increasing in all locations recently. Conclusion: HCV remains a global health concern despite tremendous progress being made. The disease burden in China improved significantly, while the burden in the United States was deteriorating, with new infections increasing recently, suggesting more targeted interventions to be established to realize the 2030 elimination goals.

Reliability and validity of the Chinese version of chronic liver disease questionnaire-nonalcoholic fatty liver disease in patients with nonalcoholic fatty liver disease: a multicenter cross-sectional survey in China.

PURPOSE: The Chronic Liver Disease Questionnaire (CLDQ)-Nonalcoholic Fatty Liver Disease (NAFLD) is a disease-specific instrument to assess the health-related quality of life (HRQL) of patients with NAFLD. In order to provide further evidence for the cross-cultural utility of this instrument in the Chinese population, we translated the CLDQ-NAFLD into Chinese and examined its reliability and validity. METHODS: Patients with NAFLD in 90 hospitals across China were enrolled in this multicenter cross-sectional survey. Eligible patients completed the Chinese version of CLDQ-NAFLD at enrollment to assess HRQL. Internal consistency of the questionnaire was assessed using Cronbach's alpha coefficient and split-half reliability. Convergent and discriminant validity were assessed using Spearman correlation coefficient. Factor analysis was used to test the construct validity. RESULTS: Between March and August 2019, 5181 patients with a mean age of 43.8 ± 13.3 years were enrolled. All domains exhibited good internal consistency, with Cronbach's alpha and split-half reliability greater than 0.70. The scaling success rate of all domains was 100% for convergent validity and 99.4% (179/180) for discriminant validity. The inter-scale correlations indicated a significant correlation between all CLDQ-NAFLD domains (r = 0.608 to 0.832, all p < 0.001). Factor analysis of 36 items extracted 6 factors, which explained 69.14% of the total variance. CONCLUSION: The Chinese version of CLDQ-NAFLD is a reliable and valid instrument for assessing the HRQL of Chinese patients with NAFLD.

Developing a New qFIBS Model Assessing Histological Features in Pediatric Patients With Non-alcoholic Steatohepatitis.

Background: The evolution of pediatric non-alcoholic fatty liver disease (NAFLD) to non-alcoholic steatohepatitis (NASH) is associated with unique histological features. Pathological evaluation of liver specimen is often hindered by observer variability and diagnostic consensus is not always attainable. We investigated whether the qFIBS technique derived from adult NASH could be applied to pediatric NASH. Materials and Methods: 102 pediatric patients (<18 years old) with liver biopsy-proven NASH were included. The liver biopsies were serially sectioned for hematoxylin-eosin and Masson trichrome staining for histological scoring, and for second harmonic generation (SHG) imaging. qFIBS-automated measure of fibrosis, inflammation, hepatocyte ballooning, and steatosis was estabilshed by using the NASH CRN scoring system as the reference standard. Results: qFIBS showed the best correlation with steatosis (r = 0.84, P < 0.001); with ability to distinguish different grades of steatosis (AUROCs 0.90 and 0.98, sensitivity 0.71 and 0.93, and specificity 0.90 and 0.90). qFIBS correlation with fibrosis (r = 0.72, P < 0.001) was good with high AUROC values [qFibrosis (AUC) > 0.85 (0.85-0.95)] and ability to distinguish different stages of fibrosis. qFIBS showed weak correlation with ballooning (r = 0.38, P = 0.028) and inflammation (r = 0.46, P = 0.005); however, it could distinguish different grades of ballooning (AUROCs 0.73, sensitivity 0.36, and specificity 0.92) and inflammation (AUROCs 0.77, sensitivity 0.83, and specificity 0.53). Conclusion: It was demonstrated that when qFIBS derived from adult NASH was performed on pediatric NASH, it could best distinguish the various histological grades of steatosis and fibrosis.

Distribution and changes in hepatitis C virus genotype in China from 2010 to 2020.

BACKGROUND: Hepatitis C virus (HCV) causes a large number of infections worldwide. New infections seem to be increasing according to a report of the World Health Organization in 2015. Although direct-acting antivirals are quite effective for most genotypes of the HCV, some genotypes fail to respond. Therefore, the trend of genotype distribution is vital to better control the development of this infection. AIM: To analyze the distribution and trends of the HCV genotype before and after the emergence of direct-acting antivirals in China. METHODS: We searched all literature published in five electronic databases-China National Knowledge Infrastructure, Wan Fang Data, VIP Chinese Journal Database, Chinese Biomedical Literature Service System, and PubMed-from January 1, 2010 to December 31, 2020. The search strategy combined medical subject headings and free-text terms, including "hepatitis C virus" or "HCV" and "genotype" or "subtype" and "China" or "Chinese". Additional relevant articles were searched by manual selection. Data were extracted to build a database. All of the data were totaled according to regions, periods, routes of transmission, and sexes. The percentages in various stratifications were calculated. RESULTS: There were 76110 samples from 30 provinces included in the study. Genotype 1 (G1) accounted for 58.2% of cases nationwide, followed by G2, G6, G3b, G3a, unclassified and mixed infections (17.5%, 7.8%, 6.4%, 4.9%, 1.8%, and 1.2%, respectively). The constitution of genotype varied among different regions, with G6 and G3b being more common in the south and southwest, respectively (28.1%, 15.4%). The past ten years have witnessed a decrease in G1 and G2 and an increase in G3 and G6 in almost all regions. The drug-use population had the most abundant genotypes, with G6 ranking first (33.3%), followed by G1 and G3b (23.4%, 18.5%). CONCLUSION: G3 and G6 pose a new challenge for HCV infection. This study revealed the distribution of HCV genotypes in China over the past 10 years, providing information for HCV management strategies.

Gadolinium-ethoxybenzyl-diethylenetriamine penta-acetic acid-enhanced magnetic resonance imaging for evaluating fibrosis regression in chronic hepatitis C patients after direct-acting antiviral.

BACKGROUND: Direct acting antiviral (DAA) therapy has enabled hepatitis C virus infection to become curable, while histological changes remain uncontained. Few valid non-invasive methods can be confirmed for use in surveillance. Gadolinium-ethoxybenzyl-diethylenetriamine penta-acetic acid (Gd-EOB-DTPA) is a liver-specific magnetic resonance imaging (MRI) contrast, related to liver function in the hepatobiliary phase (HBP). Whether Gd-EOB-DTPA-enhanced MRI can be used in the diagnosis and follow up of hepatic fibrosis in patients with chronic hepatitis C (CHC) has not been investigated. AIM: To investigate the diagnostic and follow-up values of Gd-EOB-DTPA-enhanced MRI for hepatic histology in patients with CHC. METHODS: Patients with CHC were invited to undergo Gd-EOB-DTPA-enhanced MRI and liver biopsy before treatment, and those with paired qualified MRI and liver biopsy specimens were included. Transient elastography (TE) and blood tests were also arranged. Patients treated with DAAs who achieved 24-wk sustained virological response (SVR) underwent Gd-EOB-DTPA-enhanced MRI and liver biopsy again. The signal intensity (SI) of the liver and muscle were measured in the unenhanced phase (UEP) (SIUEP-liver, SIUEP-muscle) and HBP (SIHBP-liver, SIHBP-muscle) via MRI. The contrast enhancement index (CEI) was calculated as [(SIHBP-liver/SIHBP-muscle)]/[(SIUEP-liver/SIUEP-muscle)]. Liver stiffness measurement (LSM) was confirmed with TE. Serologic markers, aspartate aminotransferase-to-platelet ratio index (APRI) and Fibrosis-4 (FIB-4), were also calculated according to blood tests. The grade of inflammation and stage of fibrosis were evaluated with the modified histology activity index (mHAI) and Ishak fibrosis score, respectively. Fibrosis regression was defined as a ≥ 1-point decrease in the Ishak fibrosis score. The correlation between the CEI and liver pathology was evaluated. The diagnostic and follow-up values of the CEI, LSM, and serologic markers were compared. RESULTS: Thirty-nine patients with CHC were enrolled [average age, 42.3 ± 14.4 years; 20/39 (51.3%) male]. Twenty-one enrolled patients had eligible paired Gd-EOB-DTPA-enhanced MRI and liver tissues after achieving SVR. The mHAI median significantly decreased after SVR [baseline 6.0 (4.5-13.5) vs SVR 2.0 (1.5-5.5), Z = 3.322, P = 0.017], but the median stage of fibrosis did not notably change (P > 0.05). Sixty pairs of qualified MRI and liver tissue samples were available for use to analyze the relationship between the CEI and hepatic pathology. The CEI was negatively correlated with the mHAI (r = -0.56, P < 0.001) and Ishak score (r = -0.69, P < 0.001). Further stratified analysis showed that the value of the CEI decreased with the progression of the stage of fibrosis rather than with the grade of necroinflammation. For patients with Ishak score ≥ 5, the areas under receiver operating characteristics curve of the CEI, LSM, APRI, and FIB-4 were approximately at baseline, 0.87-0.93, and after achieving SVR, 0.83-0.91. The CEI cut-off value was stable (baseline 1.58 and SVR 1.59), but those of the APRI (from 1.05 to 0.24), FIB-4 (from 1.78 to 1.28), and LSM (from 10.8 kpa to 7.1 kpa) decreased dramatically. The APRI and FIB-4 cannot be used as diagnostic means for SVR in patients with Ishak score ≥ 3 (P > 0.05). Seven patients achieved fibrosis regression after achieving SVR. In these patients, the CEI median increased (from 1.71 to 1.83, Z = -1.981, P = 0.048) and those of the APRI (from 1.71 to 1.83, Z = -2.878, P = 0.004) and LSM (from 6.6 to 4.8, Z = -2.366, P = 0.018) decreased. However, in patients without fibrosis regression, the medians of the APRI, FIB-4, and LSM also changed significantly (P < 0.05). CONCLUSION: Gd-EOB-DTPA-enhanced MRI has good diagnostic value for staging fibrosis in patients with CHC. It can be used for fibrotic-change monitoring post SVR in patients with CHC treated with DAAs.

Health-related quality of life in Chinese population with non-alcoholic fatty liver disease: a national multicenter survey.

BACKGROUND: Health Related Quality of Life (HRQL) is a multi-dimensional construct that can comprehensively evaluate the patient's health status, including physical, emotional, mental and social well-being. In this study, we aimed to evaluate the impact of non-alcoholic fatty liver disease (NAFLD) on HRQL in a Chinese population. METHODS: In this national multicenter cross-sectional survey, patients with NAFLD were enrolled. Chronic Liver Disease Questionnaire (CLDQ)-NAFLD was used to qualify HRQL. Univariate and multivariate analysis were used to identify independent risk factors of HRQL. RESULTS: A total of 5181 patients with NAFLD from 90 centers were enrolled in this study (mean age, 43.8 ± 13.3 years; male, 65.8%). The overall CLDQ score was 5.66 ± 0.89. Multivariate logistic regression analysis showed that body mass index (BMI: HR, 1.642; 95% CI, 1.330-2.026), alanine transaminase (ALT: HR, 1.006; 95% CI, 1.001-1.011), triglyceride (HR, 1.184; 95% CI, 1.074-1.305), disease severity (HR, 3.203; 95% CI, 1.418-7.232) and cardiovascular disease (HR, 4.305; 95% CI, 2.074-8.939) were independent risk factors for overall CLDQ score. In the logistic analyses of individual domain, BMI and triglyceride were independent risk factors of all domains. ALT, disease severity, diabetes, depression and cardiovascular disease were influencing factors for the CLDQ score of several domains. CONCLUSIONS: This national multicenter cross-sectional survey in China indicated that the HRQL in patients with NAFLD was impaired. HRQL was found to be significantly associated with sociodemographic and clinical factors. Attention should be paid to the optimally managing care of patients with NAFLD to improve their HRQL.

Histopathology and the predominantly progressive, indeterminate and predominately regressive score in hepatitis C virus patients after direct-acting antivirals therapy.

BACKGROUND: Histological changes after direct-acting antivirals (DAAs) therapy in hepatitis C virus (HCV) patients has not been elucidated. Whether the predominantly progressive, indeterminate and predominately regressive (P-I-R) score, evaluating fibrosis activity in hepatitis B virus patients has predictive value in HCV patients has not been investigated. AIM: To identify histological changes after DAAs therapy and to evaluate the predictive value of the P-I-R score in HCV patients. METHODS: Chronic HCV patients with paired liver biopsy specimens before and after DAAs treatment were included. Sustained virologic response (SVR) was defined as an undetectable serum HCV RNA level at 24 wk after treatment cessation. The Ishak system and P-I-R score were assessed. Inflammation improvement and fibrosis regression were defined as a ≥ 2-points decrease in the histology activity index (HAI) score and a ≥ 1-point decrease in the Ishak fibrosis score, respectively. Fibrosis progression was defined as a ≥ 1-point increase in the Ishak fibrosis score. Histologic improvement was defined as a ≥ 2-points decrease in the HAI score without worsening of the Ishak fibrosis score after DAAs therapy. The P-I-R score was also assessed. "absolutely reversing or advancing" was defined as the same directionality implied by both change in the Ishak score and posttreatment P-I-R score; and "probably reversing or advancing" was defined as only one parameter showing directionality. RESULTS: Thirty-eight chronic HCV patients with paired liver biopsy specimens before and after DAAs treatment were included. The mean age of these patients was 40.9 ± 14.6 years and there were 53% (20/38) males. Thirty-four percent (13/38) of patients were cirrhotic. Eighty-two percent (31/38) of patients achieved inflammation improvement. The median HAI score decreased significantly after SVR (pretreatment 7.0 vs posttreatment 2.0, Z = -5.146, P = 0.000). Thirty-seven percent (14/38) of patients achieved fibrosis improvement. The median Ishak score decreased significantly after SVR (pretreatment 4.0 vs posttreatment 3.0, Z = -2.354, P = 0.019). Eighty-two percent (31/38) of patients showed histological improvement. The P-I-R score was evaluated in 61% (23/38) of patients. The progressive group showed lower platelet (P = 0.024) and higher HAI scores (P = 0.070) before treatment. In patients with stable Ishak stage after treatment: Progressive injury was seen in 22% (4/18) of patients, 33% (6/18) were classified as indeterminate and regressive changes were seen in 44% (8/18) of patients who were judged as probably reversing by the Ishak and P-I-R systems. CONCLUSION: Significant improvement of necroinflammation and partial remission of fibrosis in HCV patients occurred shortly after DAAs therapy. The P-I-R score has potential in predicting fibrosis in HCV patients.

Impact of hepatitis C virus genotype 3 on liver disease progression in a Chinese national cohort.

BACKGROUND: Hepatitis C virus (HCV) genotype 3, particularly subtype 3b, is increasing in prevalence and distribution in China. This study evaluated the prevalence, regional distribution, clinical characteristics, host factors, treatment outcomes, and disease progression of patients with HCV genotype 3 in China. METHODS: A 5-year follow-up was preceded by a cross-sectional study. Treatment choices were at the discretion of treating physicians. Estimated infection time to overall-disease-progression (defined by ≥1 of: newly diagnosed cirrhosis; cirrhosis at baseline, Child-Turcotte-Pugh score increased 2 points or more; progression from compensated cirrhosis to decompensated cirrhosis; hepatocellular carcinoma; liver transplantation; or death) was calculated using the Kaplan-Meier method. Cox regression analyses were conducted to evaluate the risk factors for disease progression. RESULTS: The cross-sectional study enrolled 997 patients, including 91 with HCV genotype 3 infection. Among them, subtype 3b (57.1%) was more dominant than subtype 3a (38.5%). Five hundred and twelve patients were included into the follow-up phase. Among patients analyzed for estimated infection time to overall-disease-progression, 52/304 (17.1%) patients with HCV genotype 1 and 4/41 (9.8%) with HCV genotype 3 (4/26 with genotype 3b, 0/13 with genotype 3a, and 0/2 with undefined subtype of genotype 3) experienced overall-disease-progression. Patients with HCV genotype 3 were younger than those with genotype 1 (mean age: 39.5 ±â€Š8.7 vs. 46.9 ±â€Š13.6 years) and demonstrated more rapid disease progression (mean estimated infection time to overall-disease-progression 27.1 vs. 35.6 years). CONCLUSIONS: HCV genotype 3, specifically subtype 3b, is associated with more rapid progression of liver disease. Further analysis to compare HCV subtype 3a and 3b is needed in high prevalence regions. TRIAL REGISTRATION: NCT01293279, https://clinicaltrials.gov/ct2/show/NCT01293279; NCT01594554, https://clinicaltrials.gov/ct2/show/NCT01594554.

qFIBS: An Automated Technique for Quantitative Evaluation of Fibrosis, Inflammation, Ballooning, and Steatosis in Patients With Nonalcoholic Steatohepatitis.

BACKGROUND AND AIMS: Nonalcoholic steatohepatitis (NASH) is a common cause of chronic liver disease. Clinical trials use the NASH Clinical Research Network (CRN) system for semiquantitative histological assessment of disease severity. Interobserver variability may hamper histological assessment, and diagnostic consensus is not always achieved. We evaluate a second harmonic generation/two-photon excitation fluorescence (SHG/TPEF) imaging-based tool to provide an automated quantitative assessment of histological features pertinent to NASH. APPROACH AND RESULTS: Images were acquired by SHG/TPEF from 219 nonalcoholic fatty liver disease (NAFLD)/NASH liver biopsy samples from seven centers in Asia and Europe. These were used to develop and validate qFIBS, a computational algorithm that quantifies key histological features of NASH. qFIBS was developed based on in silico analysis of selected signature parameters for four cardinal histopathological features, that is, fibrosis (qFibrosis), inflammation (qInflammation), hepatocyte ballooning (qBallooning), and steatosis (qSteatosis), treating each as a continuous rather than categorical variable. Automated qFIBS analysis outputs showed strong correlation with each respective component of the NASH CRN scoring (P < 0.001; qFibrosis [r = 0.776], qInflammation [r = 0.557], qBallooning [r = 0.533], and qSteatosis [r = 0.802]) and high area under the receiver operating characteristic curve values (qFibrosis [0.870-0.951; 95% confidence interval {CI}, 0.787-1.000; P < 0.001], qInflammation [0.820-0.838; 95% CI, 0.726-0.933; P < 0.001), qBallooning [0.813-0.844; 95% CI, 0.708-0.957; P < 0.001], and qSteatosis [0.939-0.986; 95% CI, 0.867-1.000; P < 0.001]) and was able to distinguish differing grades/stages of histological disease. Performance of qFIBS was best when assessing degree of steatosis and fibrosis, but performed less well when distinguishing severe inflammation and higher ballooning grades. CONCLUSIONS: qFIBS is an automated tool that accurately quantifies the critical components of NASH histological assessment. It offers a tool that could potentially aid reproducibility and standardization of liver biopsy assessments required for NASH therapeutic clinical trials.

Risk factors for familial clustering of hepatitis C virus infection in a Chinese Han population: a cross-sectional study.

BACKGROUND: Hepatitis C is a curable disease, but reinfection from household contact may occur in patients who have achieved sustained viral response (SVR). METHODS: A total of 997 ethnic Han HCV treatment-naïve adult patients were enrolled in a cross-sectional study with stratified sampling based on the populations of five geographic regions across China to examine the genetic and physiological parameters associated with the phenomenon of HCV familial clustering. RESULTS: Of the total 997 patients, there were 59 patients who had at least one family member with HCV infection according to patient self-report. Comparison between patients with and without HCV familial clustering by univariate regression analysis showed that genotype 2, sexual transmission, long-term exposure to HCV patients, monthly family income per person less than 2000 yuan, farming occupation, and the southern and northern regions were associated with HCV familial clustering. Blood transfusion was negatively associated with HCV familial clustering. Multivariate logistic regression analysis suggested that long-term exposure to HCV patients and low family income were correlated with HCV familial clustering, whereas blood transfusion was negatively associated, which meant that blood transfusion was not the main transmission route in HCV familial clustering. CONCLUSION: Long-term exposure to HCV patients and low family income were correlated with HCV familial clustering, whereas blood transfusion was not the main transmission route in HCV familial clustering. To reduce reinfection from household contacts, education and awareness of HCV transmission routes and familial clustering should be strengthened, especially among HCV patients' family members, low-income families and non-blood transmission hepatitis C patients.

Second Harmonic Generation Reveals Subtle Fibrosis Differences in Adult and Pediatric Nonalcoholic Fatty Liver Disease.

OBJECTIVES: Investigate subtle fibrosis similarities and differences in adult and pediatric nonalcoholic fatty liver disease (NAFLD) using second harmonic generation (SHG). METHODS: SHG/two-photon excitation fluorescence imaging quantified 100 collagen parameters and determined qFibrosis values by using the nonalcoholic steatohepatitis (NASH) Clinical Research Network (CRN) scoring system in 62 adult and 36 pediatric NAFLD liver specimens. RESULTS: Six distinct parameters identified differences among the NASH CRN stages with high accuracy (area under the curve, 0835-0.982 vs 0.885-0.981, adult and pediatric). All portal region parameters showed similar changes across early stages 0, 1C, and 2, in both groups. Parameter values decreased in adults with progression from stage 1A/B to 2 in the central vein region. In children, aggregated collagen parameters decreased, but nearly all distributed collagen parameters increased from stage 1A/B to 2. CONCLUSIONS: SHG analysis accurately reproduces NASH CRN staging in NAFLD, as well as reveals differences and similarities between adult and pediatric collagen deposition not captured by currently available quantitative methods.

Automated evaluation of liver fibrosis in thioacetamide, carbon tetrachloride, and bile duct ligation rodent models using second-harmonic generation/two-photon excited fluorescence microscopy.

Animal models provide a useful platform for developing and testing new drugs to treat liver fibrosis. Accordingly, we developed a novel automated system to evaluate liver fibrosis in rodent models. This system uses second-harmonic generation (SHG)/two-photon excited fluorescence (TPEF) microscopy to assess a total of four mouse and rat models, using chemical treatment with either thioacetamide (TAA) or carbon tetrachloride (CCl4), and a surgical method, bile duct ligation (BDL). The results obtained by the new technique were compared with that using Ishak fibrosis scores and two currently used quantitative methods for determining liver fibrosis: the collagen proportionate area (CPA) and measurement of hydroxyproline (HYP) content. We show that 11 shared morphological parameters faithfully recapitulate Ishak fibrosis scores in the models, with high area under the receiver operating characteristic (ROC) curve (AUC) performance. The AUC values of 11 shared parameters were greater than that of the CPA (TAA: 0.758-0.922 vs 0.752-0.908; BDL: 0.874-0.989 vs 0.678-0.966) in the TAA mice and BDL rat models and similar to that of the CPA in the TAA rat and CCl4 mouse models. Similarly, based on the trends in these parameters at different time points, 9, 10, 7, and 2 model-specific parameters were selected for the TAA rats, TAA mice, CCl4 mice, and BDL rats, respectively. These parameters identified differences among the time points in the four models, with high AUC accuracy, and the corresponding AUC values of these parameters were greater compared with those of the CPA in the TAA rat and mouse models (rats: 0.769-0.894 vs 0.64-0.799; mice: 0.87-0.93 vs 0.739-0.836) and similar to those of the CPA in the CCl4 mouse and BDL rat models. Similarly, the AUC values of 11 shared parameters and model-specific parameters were greater than those of HYP in the TAA rats, TAA mice, and CCl4 mouse models and were similar to those of HYP in the BDL rat models. The automated evaluation system, combined with 11 shared parameters and model-specific parameters, could specifically, accurately, and quantitatively stage liver fibrosis in animal models.

Real-world treatment patterns and clinical outcomes of HCV treatment-naive patients in China: an interim analysis from the CCgenos study.

BACKGROUND AND AIM: In China, chronic hepatitis C virus (HCV) infection represents a considerable healthcare burden. Although interferon-based therapy has been the standard-of-care for many years, few long-term, real-life studies have assessed interferon-based treatment in China. The objective of CCgenos follow-up study was to analyze long-term treatment patterns and outcomes in a cohort of treatment-naïve, Han ethnic, patients with chronic HCV infection. METHODS: Patients who had participated in the CCgenos cross-sectional study were invited to enter this 5-year follow up. Clinical information and centralized HCV-RNA measures were collected at scheduled study visits every 6 months for untreated patients and every 3 months for treated patients. RESULTS: Among 512 patients enrolled, 334 (65.2%) received interferon-based treatment and 178 (34.8%) remained untreated over a median of 4.1 (1.2-4.3) years. A total of 82.8% (424/512) of patients had an IL28B CC genotype (GT); 60.7% (311/512) had HCV GT1b infection, including 121 (38.9%) untreated. Most patients with baseline cirrhosis were untreated (26/46, 56.5%). Among patients who completed treatment and 24 weeks of post-treatment follow up, the duration of interferon-based therapy was frequently longer than recommended (52.9% [92/174] of GT1b-infected were treated for > 1 year). Rates of sustained virologic response (SVR24) were 71.1% (226/318) overall; 62.4% (111/178) among patients with HCV GT1b infection; and 42.9% (15/35) among patients with cirrhosis. CONCLUSIONS: There remains a high unmet need for effective HCV treatment in China, evidenced by a high proportion of patients remaining untreated by the current standard-of-care and relatively low SVR24 rates for patients with both GT1b infection and cirrhosis.

Hepatitis B virus infection in Chinese patients with hepatitis C virus infection: prevalence, clinical characteristics, viral interactions and host genotypes: a nationwide cross-sectional study.

OBJECTIVES: Little is known about hepatitis B virus (HBV) infection in patients with hepatitis C virus (HCV) infection in China. This study aimed to evaluate the prevalence, clinical characteristics, viral interactions and host genotypes of HBV/HCV dual infection compared with HCV monoinfection. STUDY DESIGN: A cross-sectional study. SETTING: China. PARTICIPANTS AND METHODS: 997 patients with HCV from 28 university-affiliated hospitals in China were enrolled in this research. Patients were divided into two subgroups. RESULTS: The prevalence of HBV infection in patients with HCV was 4.11% (41/997). The age-specific prevalence of HBsAg was 0.70%, 3.97% and 5.85% in groups aged 18-30, 30-50 and >50 years old (p=0.057), respectively. Patients with HBV/HCV dual infection and patients with HCV monoinfection had similar HCV viral loads (5.80±0.89 vs 5.83±1.00 log10 IU/mL, p=0.904). The dominant HCV genotype was 1b in both groups (53.65% vs 56.90%, p=0.493). The protective C allele in IL-28B (rs12979860) was also the dominant allele type in both patient groups (85.36% vs 83.99%, p=0.814). Patients with HBV/HCV dual infection had a higher ratio of liver cirrhosis and hepatic decompensation than patients with HCV monoinfection (39.02% vs 17.69%, p=0.001; 31.70% vs 12.13%, p=0.001). CONCLUSIONS: The HBV burden was moderate in HCV-infected patients in China. Liver cirrhosis was more common in patients with HBV/HCV dual infection, suggesting the need for closer monitoring of dual-infected individuals. TRIAL REGISTRATION NUMBER: NCT01293279; Post-results.

CD4+CXCR5+ T cells activate CD27+IgG+ B cells via IL-21 in patients with hepatitis C virus infection.

BACKGROUND: Chronic hepatitis C virus (HCV) infection causes the skewing and activation of B cell subsets, but the characteristics of IgG+ B cells in patients with chronic hepatitis C (CHC) infection have not been thoroughly elucidated. CD4+CXCR5+ follicular helper T (Tfh) cells, via interleukin (IL)-21 secretion, activate B cells. However, the role of CD4+CXCR5+ T cells in the activation of IgG+ B cells in CHC patients is not clear. METHODS: The frequency of IgG+ B cells, including CD27-IgG+ B and CD27+IgG+ B cells, the expression of the activation markers (CD86 and CD95) in IgG+ B cells, and the percentage of circulating CD4+CXCR5+ T cells were detected by flow cytometry in CHC patients (n=70) and healthy controls (n=25). The concentrations of serum IL-21 were analyzed using ELISA. The role of CD4+CXCR5+ T cells in the activation of IgG+ B cells was investigated using a co-culture system. RESULTS: A significantly lower proportion of CD27+IgG+ B cells with increased expression of CD86 and CD95 was observed in CHC patients. The expression of CD95 was negatively correlated with the percentage of CD27+IgG+ B cells, and it contributed to CD27+IgG+ B cell apoptosis. Circulating CD4+CXCR5+ T cells and serum IL-21 were significantly increased in CHC patients. Moreover, circulating CD4+CXCR5+ T cells from CHC patients induced higher expressions of CD86 and CD95 in CD27+IgG+ B cells in a co-culture system; the blockade of the IL-21 decreased the expression levels of CD86 and CD95 in CD27+IgG+ B cells. CONCLUSIONS: HCV infection increased the frequency of CD4+CXCR5+ T cells and decreased the frequency of CD27+IgG+ B cells. CD4+CXCR5+ T cells activated CD27+IgG+ B cells via the secretion of IL-21.

Direct-acting Antiviral Agents Resistance-associated Polymorphisms in Chinese Treatment-naïve Patients Infected with Genotype 1b Hepatitis C Virus.

BACKGROUND: It has been reported that several baseline polymorphisms of direct-acting antivirals (DAAs) agents resistance-associated variants (RAVs) would affect the treatment outcomes of patients chronically infected with hepatitis C virus (CHC). The aim of this study is to investigate the prevalence of DAAs RAVs in treatment-naÏve GT1b CHC patients. METHODS: Direct sequencing and ultra-deep sequencing of the HCV NS3, NS5A, and NS5B gene were performed in baseline serum samples of treatment-naÏve patients infected with genotype 1b hepatitis C virus (HCVs). RESULTS: One hundred and sixty CHC patients were studied. Complete sequence information was obtained for 145 patients (NS3), 148 patients (NS5A), and 137 patients (NS5B). Treatment-failure associated variants of DAAs were detected: 56.6% (82/145) of the patients presented S122G for simeprevir (NS3 protease inhibitor); 10.1% (14/148) of the patients presented Y93H for daclatasvir and ledipasvir (NS5A protein inhibitors); 94.2% (129/137) of the patients presented C316N for sofosbuvir (NS5B polymerase inhibitor). Nearly, all of the DAAs RAVs detected by ultra-deep sequencing could be detected by direct sequencing. CONCLUSIONS: The majority of genotype 1b CHC patients in China present a virus population carrying HCV DAAs RAVs. Pretreatment sequencing of HCV genome might need to be performed when patients infected with GT1b HCV receiving DAAs-containing regimens in China. Population sequencing would be quite quantified for the work.

Early predictive efficacy of core antigen on antiviral outcomes in genotype 1 hepatitis C virus infected patients

Response-guided therapy is of limited use in developing countries because hepatitis C virus RNA detection by sensitive molecular methods is time- and labor-consuming and expen- sive. We evaluated early predictive efficacy of serum hepatitis C virus core antigen kinetics on sustained virologic response in patients with genotype 1 hepatitis C virus during pegylated interferon plus ribavirin treatment. For 478 patients recruited, hepatitis C virus RNAs were detected at baseline, and at weeks 4, 12, 24, 48, and 72 using Cobas TaqMan. Architect hepatitis C virus core antigen was performed at baseline, and weeks 4 and 12. Predictive values of hepatitis C virus core antigen on sustained virologic response were compared to hepatitis C virus RNA. In the first 12 weeks after treatment initiation the dynamic patterns of serum hepatitis C virus core antigen and hepatitis C virus RNA levels were similar in sustained virologic response, relapse, and null response patients groups. Although areas under the receiver operating characteristics curves of hepatitis C virus core antigen were lower than those of hepatitis C virus RNA at the same time points, modeling analysis showed that undetectable hepatitis C virus core antigen (rapid virological response based on hepatitis C virus core antigen) had similar positive predictive value on sustained virologic response to hepatitis C virus RNA at week 4 (90.4% vs 93.3%), and hepatitis C virus core antigen decrease greater than 1 log10 IU/mL (early virological response based on hepatitis C virus core antigen) had similar negative predictive value to hepatitis C virus RNA at week 12 (94.1% vs 95.Z%). Analysis on the validation group demonstrated a positive predictivevalue of 97.5% in rapid virological response based on hepatitis C virus core antigen and a negative predictive value of 100% in early virological response based on hepatitis C virus core antigen. In conclusion, hepatitis C virus core antigen is comparable to hepatitis C virus RNA in predicting sustained virologic response of chronic genotype 1 hepatitis C virus infected patients, and can be used to guide anti-hepatitis C virus treatment, especially in resource-limited areas.

Early predictive efficacy of core antigen on antiviral outcomes in genotype 1 hepatitis C virus infected patients.

Response-guided therapy is of limited use in developing countries because hepatitis C virus RNA detection by sensitive molecular methods is time- and labor-consuming and expensive. We evaluated early predictive efficacy of serum hepatitis C virus core antigen kinetics on sustained virologic response in patients with genotype 1 hepatitis C virus during pegylated interferon plus ribavirin treatment. For 478 patients recruited, hepatitis C virus RNAs were detected at baseline, and at weeks 4, 12, 24, 48, and 72 using Cobas TaqMan. Architect hepatitis C virus core antigen was performed at baseline, and weeks 4 and 12. Predictive values of hepatitis C virus core antigen on sustained virologic response were compared to hepatitis C virus RNA. In the first 12 weeks after treatment initiation the dynamic patterns of serum hepatitis C virus core antigen and hepatitis C virus RNA levels were similar in sustained virologic response, relapse, and null response patients groups. Although areas under the receiver operating characteristics curves of hepatitis C virus core antigen were lower than those of hepatitis C virus RNA at the same time points, modeling analysis showed that undetectable hepatitis C virus core antigen (rapid virological response based on hepatitis C virus core antigen) had similar positive predictive value on sustained virologic response to hepatitis C virus RNA at week 4 (90.4% vs 93.3%), and hepatitis C virus core antigen decrease greater than 1log10IU/mL (early virological response based on hepatitis C virus core antigen) had similar negative predictive value to hepatitis C virus RNA at week 12 (94.1% vs 95.2%). Analysis on the validation group demonstrated a positive predictive value of 97.5% in rapid virological response based on hepatitis C virus core antigen and a negative predictive value of 100% in early virological response based on hepatitis C virus core antigen. In conclusion, hepatitis C virus core antigen is comparable to hepatitis C virus RNA in predicting sustained virologic response of chronic genotype 1 hepatitis C virus infected patients, and can be used to guide anti-hepatitis C virus treatment, especially in resource-limited areas.