Yu-Qun Wang

Hepatocellular carcinoma associated microRNA expression signature: integrated bioinformatics analysis, experimental validation and clinical significance

microRNA (miRNA) expression profiles varied greatly among current studies due to different technological platforms and small sample size. Systematic and integrative analysis of published datesets that compared the miRNA expression profiles between hepatocellular carcinoma (HCC) tissue and paired adjacent noncancerous liver tissue was performed to determine candidate HCC associated miRNAs. Moreover, we further validated the confirmed miRNAs in a clinical setting using qRT-PCR and Tumor Cancer Genome Atlas (TCGA) dataset. A miRNA integrated-signature of 5 upregulated and 8 downregulated miRNAs was identified from 26 published datesets in HCC using robust rank aggregation method. qRT-PCR demonstrated that miR-93-5p, miR-224-5p, miR-221-3p and miR-21-5p was increased, whereas the expression of miR-214-3p, miR-199a-3p, miR-195-5p, miR-150-5p and miR-145-5p was decreased in the HCC tissues, which was also validated on TCGA dataset. A miRNA based score using LASSO regression model provided a high accuracy for identifying HCC tissue (AUC = 0.982): HCC risk score = 0.180E_miR-221 + 0.0262E_miR-21 - 0.007E_miR-223 - 0.185E_miR-130a. E_miR-n = Log 2 (expression of microRNA n). Furthermore, expression of 5 miRNAs (miR-222, miR-221, miR-21 miR-214 and miR-130a) correlated with pathological tumor grade. Cox regression analysis showed that miR-21 was related with 3-year survival (hazard ratio [HR]: 1.509, 95%CI: 1.079–2.112, P = 0.016) and 5-year survival (HR: 1.416, 95%CI: 1.057–1.897, P = 0.020). However, none of the deregulated miRNAs was related with microscopic vascular invasion. This study provides a basis for further clinical application of miRNAs in HCC.

Clinical value of integrated-signature miRNAs in colorectal cancer: miRNA expression profiling analysis and experimental validation

MicroRNA (miRNA) expression profiling of colorectal cancer (CRC) are often inconsistent among different studies. To determine candidate miRNA biomarkers for CRC, we performed an integrative analysis of miRNA expression profiling compared CRC tissues and paired neighboring noncancerous colorectal tissues. Using robust rank aggregation method, we identified a miRNA set of 10 integrated-signature miRNAs. In addition, the qRT-PCR validation demonstrated that 9 miRNAs were consistent dysregulated with the integrative analysis in CRC tissues, 4 miRNAs (miR-21-5p, miR-183-5p, miR-17-5p and miR-20a-5p) were up-regulated expression, and 5 miRNAs (miR-145-5p, miR-195-5p, miR-139-5p, miR-378a-5p and miR-143-3p) were down-regulated expression (all p < 0.05). Consistent with the initial analysis, 7 miRNAs were found to be significantly dysregulated in CRC tissues in TCGA data base, 4 miRNAs (miR-21-5p, miR-183-5p, miR-17-5p and miR-20a-5p) were significantly up-regulated expression, and 3 miRNAs (miR-145-5p, miR-139-5p and miR-378a-5p) were significantly down-regulated expression in CRC tissues (all p < 0.001). Furthermore, miR-17-5p (p = 0.011) and miR-20a-5p (p = 0.003) were up-regulated expression in the III/IV tumor stage, miR-145-5p (p = 0.028) and miR-195-5p (p = 0.001) were significantly increased expression with microscopic vascular invasion in CRC tissues, miR-17-5p (p = 0.037) and miR-145-5p (p = 0.023) were significantly increased expression with lymphovascular invasion. Moreover, Cox regression analysis of CRC patients in TCGA data base showed miR-20a-5p was correlated with survival (hazard ratio: 1.875, 95%CI: 1.088–3.232, p = 0.024). Hence, the finding of current study provides a basic implication of these miRNAs for further clinical application in CRC.

Dicer interacts with SIRT7 and regulates H3K18 deacetylation in response to DNA damaging agents.

Dicer participates in heterochromatin formation in fission yeast and plants. However, whether it has a similar role in mammals remains controversial. Here we showed that the human Dicer protein interacts with SIRT7, an NAD+-dependent H3K18Ac (acetylated lysine 18 of histone H3) deacetylase, and holds a proportion of SIRT7 in the cytoplasm. Dicer knockdown led to an increase of chromatin-associated SIRT7 and simultaneously a decrease of cytoplasmic SIRT7, while its overexpression induced SIRT7 reduction in the chromatin-associated fraction and increment in the cytoplasm. Furthermore, DNA damaging agents promoted Dicer expression, leading to decreased level of chromatin-associated SIRT7 and increased level of H3K18Ac, which can be alleviated by Dicer knockdown. Taken together with that H3K18Ac was exclusively associated with the chromatin, our findings suggest that Dicer induction by DNA damaging treatments prevents H3K18Ac deacetylation, probably by trapping more SIRT7 in the cytoplasm.

MicroRNA-581 Promotes Hepatitis B Virus Surface Antigen Expression by Targeting Dicer and EDEM1

Hepatitis B virus surface antigen (HBsAg) is an important risk factor for hepatocellular carcinoma (HCC) and is downregulated during hepatocarcinogenesis. MicroRNAs (miRNAs) are frequently deregulated in HCC tissues. However, whether the deregulation of certain miRNAs in HCC has an impact on HBsAg expression remains unclear. We found here that microRNA-581 (miR-581), which is deregulated during hepatocarcinogenesis, promoted HBsAg expression. Additionally, miR-581 targeted Dicer and endoplasmic reticulum degradation-enhancing alpha-mannosidase-like protein 1 (EDEM1) and repressed their expression. Although Dicer cannot process HBV transcripts, Dicer knockdown led to increased HBsAg secretion, most likely due to a reduction in the levels of Dicer-processed 7SL RNA fragments. Moreover, Dicer-processed 7SL RNA fragments partially inhibited the ability of miR-581 to stimulate HBsAg expression. Furthermore, we found that forced EDEM1 expression inhibited miR-581-mediated induction of HBsAg. Finally, transfection of miR-581 into HepG2.2.15 cells promoted cell proliferation and led to upregulation of genes involved in development, cell proliferation and protein secretion. Altogether, we conclude that miR-581 promotes HBsAg expression by targeting Dicer and EDEM1. Our findings suggest that downregulation of miR-581 during hepatocarcinogenesis may lead to a reduction in HBsAg expression and impede HCC development.

A nomogram for predicting prognostic value of inflammatory response biomarkers in decompensated cirrhotic patients without acute-on-chronic liver failure

Inflammation plays a vital role in liver cirrhosis progression and prognosis.

Prognostic value of DNA repair based stratification of hepatocellular carcinoma

Aberrant activation of DNA repair is frequently associated with tumor progression and response to therapy in hepatocellular carcinoma (HCC). Bioinformatics analyses of HCC data in the Cancer Genome Atlas (TCGA) were performed to define DNA repair based molecular classification that could predict the prognosis of patients with HCC. Furthermore, we tested its predictive performance in 120 independent cases. Four molecular subgroups were identified on the basis of coordinate DNA repair cluster (CDRC) comprising 15 genes in TCGA dataset. Increasing expression of CDRC genes were significantly associated with TP53 mutation. High CDRC was significantly correlated with advanced tumor grades, advanced pathological stage and increased vascular invasion rate. Multivariate Cox regression analysis indicated that the molecular subgrouping was an independent prognostic parameter for both overall survival (p = 0.004, hazard ratio (HR): 2.989) and tumor-free survival (p = 0.049, HR: 3.366) in TCGA dataset. Similar results were also obtained by analyzing the independent cohort. These data suggest that distinct dysregulation of DNA repair constituents based molecular classes in HCC would be useful for predicting prognosis and designing clinical trials for targeted therapy.

A diagnostic algorithm for assessment of liver fibrosis by liver stiffness measurement in patients with chronic hepatitis B

Steatosis could affect liver stiffness measurement in patients with nonalcoholic fatty liver disease and chronic hepatitis C. In this study, we aimed to investigate the impact of steatosis on liver stiffness in hepatitis B virus (HBV)‐infected patients and develop a diagnostic algorithm for prediction of liver fibrosis by liver stiffness based on the controlled attenuation parameter. A total of 488 HBV‐infected patients who underwent clinical examination, Fibroscan and liver biopsy were prospectively enrolled. The best liver stiffness measurement (kPa) cut‐offs for significant fibrosis (S≥3) and advanced fibrosis (S≥4) were 8.1 and 10.9, respectively. The best controlled attenuation parameter cut‐off for severe steatosis (≥30%) was 287 dB/m. Among patients with low‐grade fibrosis (S0‐S2/S0‐S3), mean liver stiffness values were significantly higher in subjects with severe steatosis or controlled attenuation parameter ≥287 dB/m compared with those without. Moreover, in subjects with low‐grade fibrosis, a higher rate of false‐positive rate was observed in patients with severe steatosis than those in patients without (F0‐F2: 28.2% vs 9.7%; F0‐F3: 17.0% vs 5.3%), and in patients with CAP≥287 dB/m compared with their counterpart (F0‐F2: 23.7% vs 9.2%; F0‐F3: 14.1% vs 4.8%). Low‐grade fibrosis was accurately identified by γ‐glutamyl transpeptidase‐to‐platelet ratio (GPR) with a cut‐off value of 0.17. In patients with GPR<0.17, similar results were observed. The presence of steatosis may lead to overestimation of fibrosis assessed by liver stiffness measurement in patient with chronic hepatitis B. A diagnostic algorithm for assessing fibrosis using liver stiffness was developed by combining both controlled attenuation parameter and GPR values.

Development and validation of a prognostic nomogram for acute-on-chronic hepatitis B liver failure

Determining individual risk of short‐term mortality in patients with acute‐on‐chronic hepatitis B liver failure (ACHBLF) is a difficult task. We aimed to develop and externally validate a prognostic nomogram for ACHBLF patients.

Decreased dicer expression enhances SRP-mediated protein targeting.

We have shown that Dicer processes 7SL RNA into different fragments ranging from ∼20 to more than 200 nucleotides. Here we addressed the molecular functions of these 7SL RNA fragments and found that some of them functioned as dominant-negative regulators of the full-length 7SL RNA, interfering with signal recognition particle (SRP) complex formation. Transfection of these 7SL RNA fragments inhibited the expression of cell surface glycoproteins, the targeting of a reporter protein to the endoplasmic reticulum, and the secretion of secreted alkaline phosphatase. These results suggest that some Dicer-processed 7SL RNA fragments interfered with SRP-mediated protein targeting. Moreover, we showed that Dicer knockdown enhanced SRP-mediated protein targeting and that transfection of a mixture of the 7SL RNA fragments partially restored this effect. Our data indicate that Dicer can fine-tune the efficiency of SRP-mediated protein targeting via processing a proportion of 7SL RNA into fragments of different lengths.

Usefulness of albumin‐bilirubin grade for evaluation of long‐term prognosis for hepatitis B‐related cirrhosis

Long‐term prognosis varies widely among patients with hepatitis B virus (HBV)‐related liver cirrhosis. Our study aimed to investigate the applicability of albumin‐bilirubin (ALBI), Child‐Pugh and model for end‐stage liver disease (MELD) scores to the long‐term prognosis prediction of HBV‐related cirrhosis. Patients diagnosed with HBV‐associated cirrhosis from the First Affiliated Hospital of Wenzhou Medical University between January 2010 and December 2015 were enrolled in this study. The patients were followed up every 3 months. The prognostic performance of ALBI in long‐term outcome prediction for HBV‐related cirrhosis was compared with Child‐Pugh and MELD scores using time‐dependent receiver operating characteristic curve (tdROC) and decision curve analysis. A total of 806 patients were included in our study with 275 (34.1%) deceased during the follow‐up. Multivariate Cox regression analysis showed that ALBI grade was an independent predictor associated with mortality. The tdROC analysis showed that ALBI score (0.787, 0.830 and 0.833) was superior to MELD (0.693, P=.003; 0.717, P<.001; 0.744, P<.001) and Child‐Pugh score (0.641, P<.001; 0.649, P<.001; 0.657, P<.001) for predicting 1‐year, 2‐year and 3‐year mortality. Additionally, decision curves also got the similar results. In addition, patients with lower ALBI score had a longer life expectancy, even among patients within the same Child‐Pugh class. Thus, ALBI score was effective in predicting the long‐term prognosis for patients with HBV‐related cirrhosis and more accurate than Child‐Pugh and MELD scores.