Azonal steatosis correlates with disease severity and poor outcome of nonalcoholic fatty liver disease.

OBJECTIVES: In this study we aimed to assess the clinicopathological characteristics and long-term prognosis of patients with nonalcoholic fatty liver disease (NAFLD) having distinct steatosis distribution patterns. METHODS: Clinicopathological data of 238 individuals with biopsy-confirmed NAFLD were collected. Nonalcoholic steatohepatitis-clinical research network (NASH-CRN) and steatosis, activity and fibrosis (SAF)/fatty liver inhibition of progression (FLIP) algorithm were used. Cumulative incidence of liver-related events (LREs) was compared by Kaplan-Meier analysis. Univariate and multivariate logistic regression analyses were used to identify independent predictors for steatosis distribution. RESULTS: Eligible patients were categorized into three groups based on their steatosis distribution, including azonal steatosis (AS) (62 [26.1%]), perivenular steatosis (PVS) (147 [61.8%]), and the pan-acinar steatosis (PAS) groups (29 [12.1%]). There were significantly higher ballooning grade and disease activity (P < 0.05), more severe fibrosis (P < 0.001), and a higher cumulative incidence of LREs (hazard ratio [HR] 8.0, 95% confidence interval [CI] 2.34-27.35, P < 0.0001) in the AS group than in the PVS and PAS groups after a median of 3.6-year follow-up. Multivariate logistic regression analysis revealed age (odds ratio [OR] 1.11, 95% CI 1.06-1.16, P < 0.001) might be independently associated with AS distribution, and PNPLA3 rs738409 CG/GG genotype (OR 3.36, 95% CI 0.98-11.47, P = 0.053) might also play a role. CONCLUSIONS: AS is associated with more severe disease activity and fibrosis stage in NAFLD, and predisposes toward poor prognosis. Age might be an independent predictor for AS in NAFLD, while PNPLA3 rs738409 CG/GG genotype might also play a role.

Histological assessment based on liver biopsy: the value and challenges in NASH drug development.

Nonalcoholic steatohepatitis (NASH) is increasingly recognized as a serious disease that can lead to cirrhosis, hepatocellular carcinoma (HCC), and death. However, there is no effective drug to thwart the progression of the disease. Development of new drugs for NASH is an urgent clinical need. Liver biopsy plays a key role in the development of new NASH drugs. Histological findings based on liver biopsy are currently used as the main inclusion criteria and the primary therapeutic endpoint in NASH clinical trials. However, there are inherent challenges in the use of liver biopsy in clinical trials, such as evaluation reliability, sampling error, and invasive nature of the procedure. In this article, we review the advantages and value of liver histopathology based on liver biopsy in clinical trials of new NASH drugs. We also discuss the challenges and limitations of liver biopsy and identify future drug development directions.

Liver stiffness measurement is a potent predictor of histological fibrosis regression after hepatitis C virus clearance.

BACKGROUND: Most of the studies on fibrosis regression prediction were based on noninvasive fibrosis markers and differ greatly. The 'Beijing fibrosis classification' can use histological results to classify fibrosis into progressive or 'nonprogressive' according to fibrotic septal morphology. We use this standard which served as the gold standard in order to find fibrosis regression predictors. AIM: To study the predictors of fibrosis regression after hepatitis C virus clearance according to histological fibrosis staging by the 'Beijing fibrosis classification'. MATERIALS AND METHODS: This was a prospective cohort study. A total of 68 patients with advanced liver fibrosis or compensated cirrhosis who achieved sustained virological response were enrolled. Patients with the Ishak scores lower than 3 seemed to have fibrosis regression. The others were divided into the fibrosis progressive group and the nonprogressive group according to the 'Beijing fibrosis classification'. Predictors of fibrosis regression were studied by logistic regression using baseline factors and the dynamic change in noninvasive fibrosis factors. RESULTS: Eighteen patients were assigned to the progressive group, and the others were assigned to the nonprogressive group. The baseline liver stiffness measurements (LSMs) of the progressive and nonprogressive groups were 14.35 (11.3, 27.3) kPa and 11.3 (8.3, 14.2) kPa, respectively, P = 0.02. The baseline LSM was the only predictor of fibrosis progression. With a cutoff of 11.85 kPa, the AUC was 0.71 (0.5, 0.9), and the negative predictive value was 0.92. CONCLUSIONS: The baseline LSM was found to be the only predictor of fibrosis regression, 11.85 kPa is a possible 'hepatic fibrosis return point'.

Regression of liver fibrosis: evidence and challenges.

It has been reported that liver fibrosis could be reversed after eliminating liver injuries. This article systematically summarizes the evidence of fibrosis regression based on histology, liver stiffness, and serum biomarkers, and discusses several clinically relevant challenges. Evidence from liver biopsy has been regarded as the gold standard in the assessment of fibrosis regression. Semi-quantitative staging and grading systems are traditionally and routinely used to define regression. Recently, the predominantly regressive, indeterminate, and predominantly progressive score was proposed, based on the regressive features from "hepatic repair complex", to provide additional information regarding the quality of fibrosis. For non-invasive assessment, although liver stiffness and serum biomarkers could be applied to reflect the dynamic changes of liver fibrosis, other confounding factors such as liver inflammation have to be considered. In conclusion, both histology and non-invasive methods can provide evidence regarding fibrosis regression. The predictive value of fibrosis regression in long-term prognosis warrants further investigation.

Non-obese patients with nonalcoholic fatty liver disease may use a lower liver stiffness cut-off to assess fibrosis stages.

OBJECTIVE: We aimed to estimate the optimal cut-off values of liver stiffness measurement (LSM) for diagnosing and staging fibrosis in non-obese and obese patients with nonalcoholic fatty liver disease (NAFLD). METHODS: NAFLD patients diagnosed by liver biopsy according to the Nonalcoholic Steatohepatitis Clinical Research Network scoring system were enrolled in this study. Non-obesity was defined as a body mass index (BMI) less than 25 kg/m2 . LSM was performed by experienced physicians within 2 weeks before or after liver biopsy. RESULTS: A total of 158 patients were included. Average BMI of the non-obese (n = 68) and obese (n = 90) groups was 23.2 ± 1.6 and 27.9 ± 2.5 kg/m2 , respectively. After adjusted for age, fibrosis stage, steatosis grade and type 2 diabetes mellitus, the obese group had a LSM of 3.522 kPa higher than the non-obese patients (P = 0.003). LSM values of the non-obese patients had a lower trend when stratified by fibrosis stage, especially in cirrhosis (F4; P = 0.021). Applying separate cut-off values for patients with NAFLD in individual fibrosis stage, 5.8 vs 7.5 kPa (≥ F1), 7.6 vs 8.5 kPa (≥ F2), 9.1 vs 11.2 kPa (≥ F3), and 12.5 vs 14.3 kPa (F4), improved their diagnostic odds ratios compared with overall cut-off values. In the non-obese NAFLD group, using a separate cut-off avoided underestimating 9.1% of patients with cirrhosis. CONCLUSIONS: Non-obese NAFLD group had lower LSM than the obese group. Different cut-off values should be used to measure liver fibrosis stage in non-obese and obese NAFLD patients.

Detection of Plasma EGFR Mutations in NSCLC Patients with a Validated ddPCR Lung cfDNA Assay.

Purpose: The clinical utility of cell-free DNA (cfDNA) to assess EGFR mutations is increasing. However, there are limited studies determining their clinical validity and utility. The value of cfDNA assays in cancer management remains controversial. Methods: In this study, we first evaluated the analytical performance of the ddPCR Lung cfDNA Assay. We next analyzed the concordance of the results with tissue amplification refractory mutation system PCR (ARMS-PCR) and plasma next-generation sequencing (NGS) genotyping. Finally, we assessed its clinical utility by exploring the association of cfDNA EGFR mutations with metastatic sites and the efficacy of EGFR-TKIs treatment. Results: The ddPCR Lung cfDNA Assay demonstrated a limit of blank of 1 droplet per reaction, an analytical specificity of 100%, and detection limit of 0.05%, 0.05%, and 0.1% for E746_A750del, L858R, and T790M, respectively. With tissue ARMS-PCR as a standard for comparison, the clinical sensitivity and specificity of ddPCR were 62.5% (15/24) and 100% (82/82) for E746_A750del, and 75.0% (15/20) and 94.2% (81/86) for L858R, respectively. The ddPCR showed high concordance with NGS in determining cfDNA EGFR mutations. Patients with bone and/or brain metastasis showed a higher detection rate and mutant abundance of cfDNA EGFR mutations compared to those with other sites of metastasis. Moreover, EGFR-TKIs treatment was effective in patients with sensitive EGFR mutations in either plasma cfDNA or tumor tissue-derived DNA. Conclusions: We validated in this study that the ddPCR Lung cfDNA Assay is reliable for detection of EGFR mutations in lung cancers, in terms of analytical performance, clinical validity and utility.

TGF-ß1 Induces the Dual Regulation of Hepatic Progenitor Cells with Both Anti- and Proliver Fibrosis.

Transforming growth factor-beta 1 (TGF-ß1) plays a central role in hepatic progenitor cells- (HPCs-) mediated liver repair and fibrosis. However, different effects of TGF-ß1 on progenitor cells have not been described. In this study, both in vitro (HPCs cocultured with hepatic stellate cells (HSCs) in transwells) and in vivo (CCl4-injured liver fibrosis rat) systems were used to evaluate the impacts. We found that HPCs pretreated with TGF-ß1 for 12 hours inhibited the activation of HSCs, while sensitization for 48 hours increased the activation of HSCs. Consistent with these in vitro results, the in vivo fibrosis rat model showed the same time-dependent dual effect of TGF-ß1. Regression of liver fibrosis as well as normalization of serum aminotransferase and albumin levels was detected in the rats transplanted with HPCs pretreated with TGF-ß1 for 12 hours. In contrast, severe liver fibrosis and elevated collagen-1 levels were detected in the rats transplanted with HPCs pretreated with TGF-ß1 for 48 hours. Furthermore, the TGF-ß1-pretreated HPCs were shown to deactivate HSCs via enhancing SERPINE1 expression. Inhibition of SERPINE1 reversed the deactivation response in a dose-dependent manner.

Oxygen-glucose deprivation regulates BACE1 expression through induction of autophagy in Neuro-2a/APP695 cells.

Our previous findings have demonstrated that autophagy regulation can alleviate the decline of learning and memory by eliminating deposition of extracellular beta-amyloid peptide (Aß) in the brain after stroke, but the exact mechanism is unclear. It is presumed that the regulation of beta-site APP-cleaving enzyme 1 (BACE1), the rate-limiting enzyme in metabolism of Aß, would be a key site. Neuro-2a/amyloid precursor protein 695 (APP695) cell models of cerebral ischemia were established by oxygen-glucose deprivation to investigate the effects of Rapamycin (an autophagy inducer) or 3-methyladenine (an autophagy inhibitor) on the expression of BACE1. Either oxygen-glucose deprivation or Rapamycin down-regulated the expression of BACE1 while 3-methyladenine up-regulated BACE1 expression. These results confirm that oxygen-glucose deprivation down-regulates BACE1 expression in Neuro-2a/APP695 cells through the introduction of autophagy.

Baseline Hepatitis B Virus DNA Level is a Promising Factor for Predicting the 3 (rd) Month Virological Response to Entecavir Therapy: A Study of Strict Defined Hepatitis B virus Induced Cirrhosis.

BACKGROUND: Cirrhosis is a common complication of chronic hepatitis B. It remains unclear if viral and biochemical parameters at baseline affect virological response to entecavir and therefore warrant investigation. In the present study, we aimed to evaluate the efficacy of entecavir therapy by monitoring virological response at the end of the 3 rd month of treatment and try to figure out whether baseline factors could help predict it in a cohort of hepatitis B virus (HBV) compensated cirrhosis patients and to determine the cut-off value of a predicting parameter. METHODS: A total of 91 nucleos(t)ide-naïve patients with HBV induced cirrhosis (compensatory stage) were enrolled in a prospective cohort. HBV DNA and alanine aminotransferase (ALT) were tested at baseline and monitored every 3-6 months after starting therapy. RESULTS: Of all 91 patients, the median follow-up time was 12 (9-24) months. Overall, 64 patients (70.3%) achieved virological response in the 3 rd month. Univariate analysis showed that the 3 rd month virological response can be predicted by baseline HBV DNA levels (P < 0.001, odds ratio [OR]: 2.13, 95% confidence interval [CI]: 1.44-3.15), ALT value (P = 0.023, OR: 1.01, 95% CI: 1.00-1.01) and hepatitis B e antigen (HBeAg) negativity (P = 0.016, OR: 0.30, 95% CI: 0.11-0.80). Multiple regression analysis showed baseline HBV DNA level was the only parameter related to full virological response. Higher baseline HBV DNA strata indicated a higher probability that HBV DNA remains detectable at the 3 rd month (P = 0.001). Area under receiver operating characteristic curve for determining the 3 rd month virological response by baseline HBV DNA was 77.6% (95% CI: 66.7-85.2%), with a best cut-off value of 5.8 log 10 . CONCLUSIONS: Baseline HBV DNA, HBeAg negativity, and ALT were independent factors contributing to virological response at the 3 rd month. Further, multiple regression showed that HBV DNA level was the only parameter predicting full virological response as early as the 3 rd month, in this cirrhosis cohort.

Increasing newly diagnosed rate and changing risk factors of HCV in Yanbian Prefecture, a high endemic area in China.

BACKGROUND: The newly diagnosed rate of HCV infection is increasing in China. However, the risk factors have not been fully identified. Here, a survey was performed in Yanbian Prefecture, a high-endemic area in China. METHODS: We identified newly diagnosed HCV infection in 2007-2011, using the local National Disease Supervision Information Management System from the Chinese Center for Disease Control and Prevention. We determined the risk factors using a case-control survey by questionnaire. RESULTS: Yanbian Prefecture had a rapid increase in the yearly newly diagnosed rate of HCV infection from 32.6 to 72.1/100.000 from the year 2007 to 2011. People aged 50-64 years had a high HCV infection of 43.4%, but only 0.3% of cases were reported in those aged less than 20 years. Cosmetic treatment, family history, blood transfusion, and dental treatment were independent risk factors for HCV infection. Unexpectedly, cosmetic treatments [odd ratio (OR) = 5.15, 95% confidence interval (CI) = 2.31-11.48, P = 0.00] and family history (OR = 4.68, 95% CI = 2.67-8.75, P = 0.00) showed a higher risk than the conventional risk factors of blood transfusion (OR = 4.49, 95% CI = 1.95-10.37, P = 0.001) and dental treatment (OR = 2.98, 95% CI = 1.42-6.25, P = 0.00). To further analyze the intrafamilial transmission, we found that spouses of HCV patients had an increased risk for acquiring HCV (OR = 5.75, 95% CI: 1.94-17.07), without significant association between either HCV RNA viral load (P = 0.29) or genotype (P = 0.43). CONCLUSIONS: HCV infection was increased in Yanbian Prefecture. Cosmetic treatment was a higher risk factor than medical procedure. HCV infection had a clear family clustering phenomenon, especially between spouses.

[An analysis of risk factors for ischemic stroke of different age and gender].

OBJECTIVE: To compare distribution difference in risk factors of patients with first-ever ischemic stroke (IS) of different age and gender. METHODS: A total of 1027 patients admitted to the neurological department in Shanghai Renji Hospital with first-ever IS were recruited and divided into young adult group (< 50 years old), middle-aged group (50 - 80 years old), and very old group (> 80 years old) according to their ages. Risk factor analysis included history of smoking, high alcohol consumption, hypertension (HT), diabetes mellitus (DM), heart diseases, atrial fibrillation (AF) and family history of cardiovascular diseases. RESULTS: Female patients were globally older than male patients (71.1 vs 65.7, P < 0.001) at the first attack of IS and having higher prevalence of DM (26.8% vs 19.2%, P = 0.004), heart diseases (28.8% vs 19.2%, P < 0.001) and AF (7.6% vs 3.9%, P = 0.009). However, female patients were less likely to drink heavily (1.0% vs 31.6%, P < 0.001) or smoke (4.4% vs 59.9%, P < 0.001) than the male patients. The rates of smoking and heavy drinking in young adult group were higher than that in other two groups. Patients in very old group had higher prevalence of heart diseases and AF but lower proportion of positive family cardiovascular diseases history than patients in other two groups. HT and DM were equally frequent among three groups. In young adult group, female patients were more likely to have heart diseases and family history of heart diseases (P = 0.015 and P = 0.048). In middle-old group, HT, DM, heart disease and AF were more common in women than in men (P = 0.021, P = 0.004, P = 0.001 and P = 0.039). CONCLUSION: There are differences in risk factor distribution in patients with first-ever IS of different age and gender. Therefore, screening and health education should be performed in allusion to different risk factors.

Age- and gender-specific prevalence of risk factors in patients with first-ever ischemic stroke in china.

Background. Evidences are accumulating that age and gender have great impact on the distribution of stroke risk factors. Such data are lacking in Chinese population. Methods. 1027 patients with first-ever ischemic stroke (IS) were recruited and divided into young adult (<50 years), middle-aged (50∼80 years), and very old (>80 years) groups according to stroke onset ages. Vascular risk factors were collected and compared among groups. Results. Female patients were globally older than male patients at stroke onset and having higher prevalence of diabetes mellitus (DM), heart diseases, and atrial fibrillation (AF). However, females were less likely to drink heavily or smoke than males. Young patients had a much higher proportion of smoking and drinking than middle-aged and very old patients and the highest family history of hypertension, while very old patients had the highest prevalence of heart diseases and AF but lowest proportion of positive family history of vascular diseases. Hypertension and DM were equally frequent among three groups. Conclusion. Our study showed that vascular risk factors had a specific age and gender distribution pattern in Chinese IS patients. Secondary prevention strategy should emphasize on the control of different risk factors based on patient's age and gender.

Expressions of putative cancer stem cell markers ABCB1, ABCG2, and CD133 are correlated with the degree of differentiation of gastric cancer.

BACKGROUND: The present study was carried out to determine whether a quantitative relationship exists between the expressions of 3 cancer stem cell (CSC) markers and the degree of differentiation of gastric cancer. METHODS: The expressions of 3 putative CSC markers, ABCB1, ABCG2, and CD133, were detected in 90 human gastric adenocarcinoma cases by immunofluorescence assay. The differentiation statuses of 3 gastric cancer cell lines (the undifferentiated gastric cancer cell line HGC-27, the poorly differentiated gastric cancer cell line BGC-823, and the moderately-poorly differentiated gastric adenocarcinoma cell line SGC-7901) were observed and compared by performing the 3-[4,5-dimethylthiazol-2yl]-2,5-diphenyl tetrazolium bromide (MTT) assay. Gastric xenotransplant cancers in nude mice were constructed to compare the malignancy of the 3 variously differentiated gastric cancer cell lines. The expressions of the 3 putative CSC markers were also detected in the 3 gastric cancer cell lines in vitro by flow cytometric analysis and in the 3 gastric xenotransplant cancers in vivo by immunofluorescence staining. RESULTS: The expressions of ABCB1, ABCG2, and CD133 were generally correlated with the degree of differentiation of the gastric cancers. In the human gastric adenocarcinomas and in the cancer cell lines, the expressions of ABCB1, ABCG2, and CD133 increased with the increases in the malignancy grades of the gastric cancers. In the human gastric adenocarcinomas, poorly differentiated adenocarcinoma expressed more ABCB1, ABCG2, and CD133 than well-differentiated adenocarcinoma. In addition, the expressions of ABCB1 and CD133 were higher in the diffuse type than in the intestinal type of human gastric cancers. The undifferentiated cell line HGC-27 expressed more putative CSC markers than the moderately-poorly differentiated cell line SGC-7901. Similar results were observed in the xenotransplant tumors that arose from the 3 gastric cancer cell lines. CONCLUSIONS: The expressions of the CSC markers ABCB1, ABCG2, and CD133 differed in the gastric cancers with various degrees of differentiation, with poorly differentiated gastric cancer expressing relatively more CSC markers.