Abnormal phosphorylation / dephosphorylation and Ca2+ dysfunction in heart failure.

Heart failure (HF) can be caused by a variety of causes characterized by abnormal myocardial systole and diastole. Ca2+ current through the L-type calcium channel (LTCC) on the membrane is the initial trigger signal for a cardiac cycle. Declined systole and diastole in HF are associated with dysfunction of myocardial Ca2+ function. This disorder can be correlated with unbalanced levels of phosphorylation / dephosphorylation of LTCC, endoplasmic reticulum (ER), and myofilament. Kinase and phosphatase activity changes along with HF progress, resulting in phased changes in the degree of phosphorylation / dephosphorylation. It is important to realize the phosphorylation / dephosphorylation differences between a normal and a failing heart. This review focuses on phosphorylation / dephosphorylation changes in the progression of HF and summarizes the effects of phosphorylation / dephosphorylation of LTCC, ER function, and myofilament function in normal conditions and HF based on previous experiments and clinical research. Also, we summarize current therapeutic methods based on abnormal phosphorylation / dephosphorylation and clarify potential therapeutic directions.

Hypermethylation of the glutathione peroxidase 4 gene promoter is associated with the occurrence of immune tolerance phase in chronic hepatitis B.

BACKGROUND: Hepatitis B virus (HBV) infection is a public health problem that seriously threatens human health. This study aimed to investigate the clinical significance of glutathione peroxidase 4(GPX4) in the occurrence and development of chronic hepatitis B (CHB). METHODS: A total of 169 participants including 137 patients with CHB and 32 healthy controls (HCs) were recruited. We detected the expression of GPX4 and stimulator of interferon genes (STING) in peripheral blood mononuclear cells (PBMCs) by real-time quantitative polymerase chain reaction (RT-qPCR). The methylation level of GPX4 gene promoter in PBMCs was detected by TaqMan probe-based quantitative methylation-specific PCR (MethyLight). Enzyme-linked immunosorbent assay (ELISA) was performed to detect the serum levels of GPX4, IFN-ß, oxidative stress (OS) related molecules, and pro-inflammatory cytokines. RESULTS: The expression levels of GPX4 in PBMCs and serum of CHB patients were lower than those of HCs, but the methylation levels of GPX4 promoter were higher than those of HCs, especially in patients at the immune tolerance phase. STING mRNA expression levels in PBMCs and serum IFN-ß levels of patients at the immune activation phase and reactivation phase of CHB were higher than those at other clinical phases of CHB and HCs. GPX4 mRNA expression level and methylation level in PBMCs from patients with CHB had a certain correlation with STING and IFN-ß expression levels. In addition, the methylation level of the GPX4 promoter in PBMCs from patients with CHB was correlated with molecules associated with OS and inflammation. CONCLUSIONS: GPX4 may play an important role in the pathogenesis and immune tolerance of CHB, which may provide new ideas for the functional cure of CHB.

Biological aging mediates the associations of metabolic score for insulin resistance with all-cause and cardiovascular disease mortality among US adults: A nationwide cohort study.

AIM: To investigate the associations of metabolic score for insulin resistance (METS-IR) with all-cause and cardiovascular disease (CVD)-specific mortality and the potential mediating role of biological ageing. METHODS: A cohort of 19 204 participants from the National Health and Nutrition Examination Survey (NHANES) 1999-2018 was recruited for this study. Cox regression models, restricted cubic splines, and Kaplan-Meier survival curves were used to determine the relationships of METS-IR with all-cause and CVD-specific mortality. Mediation analyses were performed to explore the possible intermediary role of biological ageing markers, including phenotypic age (PhenoAge) and biological age (BioAge). RESULTS: During a median follow-up of 9.17 years, we observed 2818 deaths, of which 875 were CVD-specific. Multivariable Cox regression showed that the highest METS-IR level (Q4) was associated with increased all-cause (hazard ratio [HR] 1.38, 95% confidence interval [CI] 1.14-1.67) and CVD mortality (HR 1.52, 95% CI 1.10-2.12) compared with the Q1 level. Restricted cubic splines showed a nonlinear relationship between METS-IR and all-cause mortality. Only METS-IR above the threshold (41.02 µg/L) was positively correlated with all-cause death. METS-IR had a linear positive relationship with CVD mortality. In mediation analyses, we found that PhenoAge mediated 51.32% (p < 0.001) and 41.77% (p < 0.001) of the association between METS-IR and all-cause and CVD-specific mortality, respectively. For BioAge, the mediating proportions of PhenoAge were 21.33% (p < 0.001) and 15.88% (p < 0.001), respectively. CONCLUSIONS: This study highlights the detrimental effects of insulin resistance, as measured by METS-IR, on all-cause and CVD mortality. Moreover, it underscores the role of biological ageing in mediating these associations, emphasizing the need for interventions targeting both insulin resistance and ageing processes to mitigate mortality risks in metabolic disorders.

Development of an advanced separation and characterization platform for mRNA and lipid nanoparticles using multi-detector asymmetrical flow field-flow fractionation.

In recent years, the use of lipid nanoparticles (LNPs) for delivery of messenger RNA (mRNA)-based therapies has gained substantial attention in the field of drug development. In such an application, multiple LNP attributes have to be carefully characterized to ensure product safety and quality, whereas accurate and efficient characterization of these complex mRNA-LNP formulations remains a challenging endeavor. Here, we present the development and application of an online separation and characterization platform designed for the isolation and in-depth analysis of mRNAs and mRNA-loaded LNPs. Our asymmetrical flow field-flow fractionation with a multi-detector (MD-AF4) method has demonstrated exceptional resolution between mRNA-LNPs and mRNAs, delivering excellent recoveries (over 70%) for both analytes and exceptional repeatability. Notably, this platform allows for comprehensive and multi-attribute LNP characterization, including online particle sizing, morphology characterization, and determination of encapsulation efficiency, all within a single injection. Furthermore, real-time online sizing by synchronizing multi-angle light scattering (MALS) and dynamic light scattering (DLS) presented higher resolution over traditional batch-mode DLS, particularly in differentiating heterogeneous samples with a low abundance of large-sized particles. Additionally, our method proves to be a valuable tool for monitoring LNP stability under varying stress conditions. Our work introduces a robust and versatile analytical platform using MD-AF4 that not only efficiently provides multi-attribute characterizations of mRNA-LNPs but also holds promise in advancing studies related to formulation screening, quality control, and stability assessment in the evolving field of nanoparticle delivery systems for mRNAs.

How do taxi drivers expose to fine particulate matter (PM2.5) in a Chinese megacity: a rapid assessment incorporating with satellite-derived information and urban mobility data.

BACKGROUND: Taxi drivers in a Chinese megacity are frequently exposed to traffic-related particulate matter (PM2.5) due to their job nature, busy road traffic, and urban density. A robust method to quantify dynamic population exposure to PM2.5 among taxi drivers is important for occupational risk prevention, however, it is limited by data availability. METHODS: This study proposed a rapid assessment of dynamic exposure to PM2.5 among drivers based on satellite-derived information, air quality data from monitoring stations, and GPS-based taxi trajectory data. An empirical study was conducted in Wuhan, China, to examine spatial and temporal variability of dynamic exposure and compare whether drivers' exposure exceeded the World Health Organization (WHO) and China air quality guideline thresholds. Kernel density estimation was conducted to further explore the relationship between dynamic exposure and taxi drivers' activities. RESULTS: The taxi drivers' weekday and weekend 24-h PM2.5 exposure was 83.60 µg/m3 and 55.62 µg/m3 respectively, 3.4 and 2.2 times than the WHO's recommended level of 25 µg/m3. Specifically, drivers with high PM2.5 exposure had a higher average trip distance and smaller activity areas. Although major transportation interchanges/terminals were the common activity hotspots for both taxi drivers with high and low exposure, activity hotspots of drivers with high exposure were mainly located in busy riverside commercial areas within historic and central districts bounded by the "Inner Ring Road", while hotspots of drivers with low exposure were new commercial areas in the extended urbanized area bounded by the "Third Ring Road". CONCLUSION: These findings emphasized the need for air quality management and community planning to mitigate the potential health risks of taxi drivers.

Hypomethylation of glycine dehydrogenase promoter in peripheral blood mononuclear cells is a new diagnostic marker of hepatitis B virus-associated hepatocellular carcinoma.

BACKGROUND: Glycine dehydrogenase (GLDC) plays an important role in the initiation and proliferation of several human cancers. In this study, we aimed to detect the methylation status of GLDC promoter and its diagnostic value for hepatitis B virus-associated hepatocellular carcinoma (HBV-HCC). METHODS: We enrolled 197 patients, 111 with HBV-HCC, 51 with chronic hepatitis B (CHB), and 35 healthy controls (HCs). The methylation status of GLDC promoter in peripheral mononuclear cells (PBMCs) was identified by methylation specific polymerase chain reaction (MSP). The mRNA expression was examined using real-time quantitative polymerase chain reaction (qPCR). RESULTS: The methylation frequency of the GLDC promoter was significantly lower in HBV-HCC patients (27.0%) compared to that in CHB patients (68.6%) and HCs (74.3%) (P < 0.001). The methylated group had lower alanine aminotransferase level (P = 0.035) and lower rates of tumor node metastasis (TNM) III/IV (P = 0.043) and T3/T4 (P = 0.026). TNM stage was identified to be an independent factor for GLDC promoter methylation. GLDC mRNA levels in CHB patients and HCs were significantly lower than those in HBV-HCC patients (P = 0.022 and P < 0.001, respectively). GLDC mRNA levels were significantly higher in HBV-HCC patients with unmethylated GLDC promoters than those with methylated GLDC promoters (P = 0.003). The diagnostic accuracy of alpha-fetoprotein (AFP) combined with GLDC promoter methylation for HBV-HCC was improved compared with that of AFP alone (AUC: 0.782 vs. 0.630, P < 0.001). In addition, GLDC promoter methylation was an independent predictor for overall survival of HBV-HCC patients (P = 0.038). CONCLUSIONS: The methylation frequency of GLDC promoter was lower in PBMCs from HBV-HCC patients than that from patients with CHB and HCs. The combination of AFP and GLDC promoter hypomethylation significantly improved the diagnostic accuracy of HBV-HCC.

IL-22-producing CD3 + CD8- T cells increase in immune clearance stage of chronic HBV infection and correlate with the response of Peg-interferon treatment.

Interleukin (IL)-22 regulates host defense. This study investigated the predominant IL-22-producing cell subsets under HBV associated immune stages. We found circulating IL-22-producing CD3 + CD8- T cells were significantly increased in immune active (IA) stage than those in immunotolerant stage, inactive carrier and healthy controls (HCs). The plasma IL-22 level was higher in IA and HBeAg-negative CHB compared to HCs. Importantly, CD3 + CD8- T cells were identified as the predominant source of plasma IL-22 production. Up-regulated IL-22-producing CD3 + CD8- T cells obviously correlated with the grade of intrahepatic inflammation. The proportions of IL-22-producing CD3 + CD8- T cells were significantly down-regulated after 48 weeks of Peg-interferon treatment, and the differences were of great significance in patients with normalize ALT levels at 48 weeks, rather than those with elevated ALT levels. In conclusion, IL-22 might play a proinflammatory function in. chronic HBV infected patients with active inflammation and Peg-interferon treatment could attenuate the degree of liver inflammation through down-regulating IL-22-producing CD3 + CD8- T cells.

MicroRNA-210-3p mediates trabecular meshwork extracellular matrix accumulation and ocular hypertension - Implication for novel glaucoma therapy.

Elevation of intraocular pressure (IOP) is a major, controllable risk factor of primary open-angle glaucoma (POAG). Transforming growth factor-ß2 (TGF-ß2)-induced excessive accumulation of extracellular matrix (ECM) in the trabecular meshwork (TM) has been demonstrated to contribute significantly to the development of high IOP. We previously showed that treatment with salidroside (Sal), a plant-derived glucoside, can ameliorate the TGF-ß2-induced ECM expression in cultured human TM cells and reduce TGF-ß2-induced ocular hypertension in mice. In the current study, its underlying molecular mechanism associated with microRNA-210-3p (miR-210-3p) was characterized. We discovered that, in TM tissues of POAG patients, there was an increase in miR-210-3p. And miR-210-3p mediated a portion of the pathological effects of TGF-ß2 in vitro (excessive accumulation of ECM in cultured human TM cells) and in vivo (mouse ocular hypertension and ECM accumulation in the TM). Most interestingly, miR-210-3p was down-regulated by Sal, which appeared to mediate a significant portion of its IOP-lowering effect. Thus, these results shed light on the probable molecular mechanisms of TGF-ß2 and Sal and indicate that manipulation of miR-210-3p level/activity represents a potential new therapeutic strategy for POAG.

Hypermethylation of thymosin ß4 predicts a poor prognosis for patients with acute-on-chronic hepatitis B liver failure.

BACKGROUND: It has been demonstrated that thymosin ß4 (Tß4) could inflect the severity of acute-on-chronic hepatitis B liver failure (ACHBLF), but the relationship between its methylation status and the prognosis of liver failure is not clear. This study aimed to determine Tß4 promoter methylation status in patients with ACHBLF and to evaluate its prognostic value. METHODS: The study recruited 115 patients with ACHBLF, 80 with acute-on-chronic hepatitis B pre-liver failure (pre-ACHBLF), and 86 with chronic hepatitis B (CHB). In addition, there were 36 healthy controls (HCs) from the Department of Hepatology, Qilu Hospital of Shandong University. The 115 patients with ACHBLF were divided into three subgroups: 33 with early stage ACHBLF (E-ACHBLF), 42 with mid-stage ACHBLF (M-ACHBLF), and 40 with advanced stage ACHBLF (A-ACHBLF). Tß4 promoter methylation status in peripheral blood mononuclear cells (PBMCs) was measured by methylation-specific polymerase chain reaction, and mRNA was detected by quantitative real-time polymerase chain reaction. RESULTS: Methylation frequency of Tß4 was significantly higher in patients with ACHBLF than in those with pre-ACHBLF, CHB or HCs. However, expression of Tß4 mRNA showed the opposite trend. In patients with ACHBLF, Tß4 promoter methylation status correlated negatively with mRNA levels. The 3-month mortality of ACHBLF in the methylated group was significantly higher than that in the unmethylated group. Also, Tß4 promoter methylation frequency was lower in survivors than in non-survivors. When used to predict the 1-, 2-, and 3-month incidence of ACHBLF, Tß4 methylation status was better than the model for end-stage liver disease (MELD) score. The predictive value of Tß4 methylation was higher than that of MELD score for the mortality of patients with E-ACHBLF and M-ACHBLF, but not for A-ACHBLF. CONCLUSIONS: Tß4 methylation might be an important early marker for predicting disease incidence and prognosis in patients with ACHBLF.

Coupling Benzylamine Oxidation with CO2 Photoconversion to Ethanol over a Black Phosphorus and Bismuth Tungstate S-Scheme Heterojunction.

Photoconversion of CO2 and H2 O into ethanol is an ideal strategy to achieve carbon neutrality. However, the production of ethanol with high activity and selectivity is challenging owing to the less efficient reduction half-reaction involving multi-step proton-coupled electron transfer (PCET), a slow C-C coupling process, and sluggish water oxidation half-reaction. Herein, a two-dimensional/two-dimensional (2D/2D) S-scheme heterojunction consisting of black phosphorus and Bi2 WO6 (BP/BWO) was constructed for photocatalytic CO2 reduction coupling with benzylamine (BA) oxidation. The as-prepared BP/BWO catalyst exhibits a superior photocatalytic performance toward CO2 reduction, with a yield of 61.3 µmol g-1 h-1 for ethanol (selectivity of 91 %).In situ spectroscopic studies and theoretical calculations reveal that S-scheme heterojunction can effectively promote photogenerated carrier separation via the Bi-O-P bridge to accelerate the PCET process. Meanwhile, electron-rich BP acts as the active site and plays a vital role in the process of C-C coupling. In addition, the substitution of BA oxidation for H2 O oxidation can further enhance the photocatalytic performance of CO2 reduction to C2 H5 OH. This work opens a new horizon for exploring novel heterogeneous photocatalysts in CO2 photoconversion to C2 H5 OH based on cooperative photoredox systems.

Spectroscopy-Based Local Modeling Method for High-Throughput Quantification of Nucleic Acid Loading in Lipid Nanoparticles.

Lipid nanoparticles (LNPs) are the most widely investigated delivery systems for nucleic acid-based therapeutics and vaccines. Loading efficiency of nucleic acids may vary with formulation conditions, and it is considered one of the critical quality attributes of LNP products. Current analytical methods for quantification of cargo loading in LNPs often require external standard preparations and preseparation of unloaded nucleic acids from LNPs; therefore, they are subject to tedious and lengthy procedures, LNP stability, and unpredictable recovery rates of the separated analytes. Here, we developed a modeling approach, which was based on locally weighted regression (LWR) of ultraviolet (UV) spectra of unpurified samples, to quantify the loading of nucleic acid cargos in LNPs in-situ. We trained the model to automatically tune the training library space according to the spectral features of a query sample so as to robustly predict the nucleic acid cargo concentration and rank loading capacity with similar performance as the more complicated experimental approaches. Furthermore, we successfully applied the model to a wide range of nucleic acid cargo species, including antisense oligonucleotides, single-guided RNA, and messenger RNA, in varied lipid matrices. The LWR modeling approach significantly saved analytical time and efforts by facile UV scans of 96-well sample plates within a few minutes and with minimal sample preprocessing. Our proof-of-concept study presented the very first data mining and modeling strategy to quantify nucleic acid loading in LNPs and is expected to better serve high-throughput screening workflows, thereby facilitates early-stage optimization and development of LNP formulations.

Prognostic potential of the small GTPase Ran and its methylation in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is a common malignant tumor with high mortality. The prognostic significance of Ran, a member of Ras superfamily, remains unclear in HCC patients. METHODS: Based on The Cancer Genome Atlas (TCGA) database and Tumor Immune Estimation Resource (TIMER), we analyzed the correlations among Ran expression, promoter methylation and immune cell infiltration. We also investigated the Ran expression levels in HCC tissues and normal tissues by using quantitative real-time PCR. RESULTS: Ran mRNA expression was significantly increased in HCC tissues compared with the normal tissues (P < 0.001). Time-dependent receiver operating characteristic (ROC) curves showed that Ran expression had predictive value of the 1-, 3- and 5-year overall survival for HCC patients, and the areas under the curves (AUC) were 0.747, 0.634 and 0.704, respectively. Cox regression analysis showed that Ran expression was an independent prognostic factor for HCC patients (HR = 1.492, 95% CI: 1.129-1.971, P = 0.005). We also found a negative relationship between Ran mRNA expression and its promoter methylation (r = -0.36, P < 0.001). High Ran expression and promoter hypomethylation predicted worse overall survival and progression-free survival (P < 0.05) and were involved in the progression of HCC. Ran expression exhibited significant correlations with immune infiltrates and prognostic immune-related genes. CONCLUSIONS: The present study provides further insight into the prognosis of HCC, and Ran could serve as a biomarker for predicting the survival of HCC patients.

A Machine Vision-Based Pipe Leakage Detection System for Automated Power Plant Maintenance.

Industrial pipework maintenance inspection can be automated through machine vision-based effusion monitoring. However, colorless effusions such as water can be difficult to detect in a complex industrial environment due to weak illumination and poor visibility of the background. This paper deploys the reflective characteristics of effusion and its lower temperature compared to the environment in order to develop an automatic inspection system for power plant pipeworks' maintenance. Such a system is aimed at detecting the colorless fluid effusion based on dual source images and a contour features algorithm. In this respect, a visible light source unit highlights the reflective features of the effusion edge. Meanwhile, high-definition images of the potential effusion are acquired under both visible and infrared lights. A customized image processing procedure extracts the potential effusion features from the infrared image to retrieve the region of interest for segmentation purposes and transfer such information to the visible light image to determine the effusion contour. Finally, a decision-making support tool based on the image contour closure is enabled for classification purposes. The implementation of the proposed system is tested on a real industrial environment. Experimental results show a classification accuracy up to 99%, demonstrating excellent suitability in meeting industrial requirements.

Hypermethylation of Cyclin D2 Predicts Poor Prognosis of Hepatitis B Virus-Associated Hepatocellular Carcinoma after Hepatectomy.

Prognosis of patients with hepatocellular carcinoma remains poor because of progression of hepatocellular carcinoma and high recurrence rates. Cyclin D2 (CCND2) plays a vital role in regulating the cell cycle; indeed, aberrant methylation of CCND2 is involved in the development of hepatocellular carcinoma. Therefore, we aimed to investigate levels of CCND2 methylation in patients with hepatitis B virus (HBV)-associated hepatocellular carcinoma and to evaluate its prognostic significance after hepatectomy. In total, 257 subjects were enrolled (166 hepatocellular carcinoma patients undergoing surgical resection, 61 chronic hepatitis B (CHB) patients, and 30 healthy controls). CCND2 methylation in peripheral blood mononuclear cells was measured quantitatively using MethyLight. We found that CCND2 methylation levels in patients with HBV-associated hepatocellular carcinoma were significantly higher than in CHB patients (P < 0.001) or healthy controls (P < 0.001). Within the hepatocellular carcinoma group, CCND2 methylation levels were higher in patients with portal vein invasion, early tumor recurrence, TNM III/IV stage, and tumor size ≥ 5 cm (P < 0.05). Furthermore, higher levels of CCND2 methylation were associated with worse overall survival and disease-free survival (P = 0.005 and P < 0.001, respectively). Multivariate analysis identified CCND2 methylation as an independent prognostic factor for early tumor recurrence (P = 0.021), overall survival (P = 0.022), and disease-free survival (P < 0.001) in hepatocellular carcinoma patients after resection. In conclusion, hypermethylation of CCND2 may have clinical utility for predicting a high risk of poor prognosis and early tumor recurrence in patients with HBV-associated hepatocellular carcinoma after hepatectomy.

A more secure and effective method for throat swab collection: The importance of adequate exposure of oral cavity in COVID-19 specimen collection.

OBJECTIVES: This study aims to propose a novel and effective throat swab collection method for coronavirus disease 2019 (COVID-19). METHODS: The subjects were randomly divided into two groups. The subjects were asked to open their mouth to make "ah" sound (traditional method) or simulate yawn (improved method) for throat swab collection. The usage of tongue depressor, collection time, adverse reactions and subjective discomfort (VAS score) were compared. The collection time, comprehensive indicators of adverse reactions and VAS score were also compared among three collectors. RESULTS: The tongue depressor was less used in the improved group (χ2 = 40.186, P < 0.01). The average collection time of the traditional group was 5.44 ± 2.97 and that of the improved group was 4.00 ± 2.31 (P < 0.01). The subjects in the improved group had fewer and milder adverse reactions. The VAS score of subjects in the improved group was lower than that in the traditional group (P < 0.01). Among different collectors, the collection time, comprehensive indicators of adverse reactions and VAS were the same as the overall trend. CONCLUSION: Simulating yawn is a safer and faster throat swab collection method.

Combined use of murine double minute-2 promoter methylation and serum AFP improves diagnostic efficiency in hepatitis B virus-related hepatocellular carcinoma.

Objective: Hepatocellular carcinoma (HCC) accounts for approximately 85% of all cases of liver cancer. In China, chronic hepatitis B virus-related HCC (HBV-related HCC) is the most common type of HCC. However, the majority of HBV-related HCC patients are asymptomatic, and the best opportunities for treating these patients are missed. The precise diagnosis of HBV-related HCC is crucial. The main purpose of this study was to evaluate the diagnostic value of murine double minute-2 (MDM2) promoter methylation in HBV-related HCC patients. Methods: The methylation status of the MDM2 promoter was detected by methylation-specific PCR. The MDM2 expression levels were validated by quantitative real-time PCR. Enzyme-linked immunosorbent assay was used to determine the levels of interleukin-6 (IL-6) and tumor-necrosis factor-α (TNF-α) in plasma. Results: The methylation frequency of the MDM2 promoter was decreased in HBV-related HCC patients. The MDM2 mRNA levels of patients with HBV-related HCC were higher than those of patients with liver cirrhosis and chronic hepatitis B. The plasma levels of IL-6 and TNF-α were significantly higher in HBV-related HCC patients than that in liver cirrhosis and chronic hepatitis B patients. The TNF-α levels were higher in the unmethylated MDM2 promoter group than in the methylated MDM2 promoter group in HBV-related HCC patients. Moreover, the combination of MDM2 promoter methylation and alpha-fetoprotein (AFP) improved the diagnosis of HBV-related HCC. Conclusions: Our study indicates, for the first time, that MDM2 promoter hypomethylation is present in HBV-related HCC patients. The combination of MDM2 promoter methylation and AFP can greatly improve diagnostic efficiency in HBV-related HCC, which might provide a new method for HBV-related HCC diagnosis.

Measurement of Cyclin D2 (CCND2) Gene Promoter Methylation in Plasma and Peripheral Blood Mononuclear Cells and Alpha-Fetoprotein Levels in Patients with Hepatitis B Virus-Associated Hepatocellular Carcinoma.

BACKGROUND Alpha-fetoprotein (AFP) is widely used to screen for hepatocellular carcinoma (HCC). However, the use of this biomarker has been challenged due to its low sensitivity and high rate of false negatives. In this study, we evaluated the diagnostic capability of cyclin D2 (CCND2) promoter methylation in patients with HCC related to hepatitis B virus (HBV). MATERIAL AND METHODS Using methylation-specific PCR and quantitative real-time PCR, we measured methylation status and mRNA levels of CCND2 in plasma and peripheral blood mononuclear cells (PBMCs) from 275 subjects: 75 patients with chronic hepatitis B (CHB), 47 with liver cirrhosis (LC), 118 with HCC, and 35 healthy controls (HCs). RESULTS The methylation rate of the CCND2 promoter was significantly higher in HCC patients than in patients without HCC (P<0.001). Furthermore, advanced HCC (TNM III/IV) was associated with a significantly higher frequency of CCND2 methylation and lower CCND2 mRNA levels than early-stage disease (TNM I/II; P<0.05). Combined measurement of CCND2 methylation and AFP yielded significantly higher sensitivity and area under the curve (AUC) than AFP alone in distinguishing patients with HCC from subjects with LC and CHB (P<0.001). CONCLUSIONS CCND2 methylation may be useful for predicting HCC progression. In addition, combined measurement of CCND2 methylation and AFP could serve as a non-invasive diagnostic marker for patients with HBV-related HCC.

Diagnostic Value of the Hypomethylation of the WISP1 Promoter in Patients with Hepatocellular Carcinoma Associated with Hepatitis B Virus.

Wnt1-inducible signaling pathway protein 1 (WISP1) regulates cell proliferation, differentiation, adhesion, migration and survival. Abnormal WISP1 expression is associated with the carcinogenesis of hepatocellular carcinoma (HCC). Aberrant DNA methylation is one of the major epigenetic alterations in HCC. However, the methylation status of the WISP1 promoter is still unclear. We therefore aimed to determine the methylation status of the WISP1 promoter and evaluate its clinical value in HCC. The study enrolled 251 participants, including 123 participants with HCC, 90 participants with chronic hepatitis B (CHB) and 38 healthy controls (HCs). WISP1 methylation status, mRNA levels and plasma soluble WISP1 were detected by methylation-specific polymerase chain reaction (MSP), quantitative real-time PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA), respectively. We found that the methylation frequency of WISP1 in patients with HCC was significantly lower than that in patients with CHB and HCs, while the relative expression levels of WISP1 mRNA were markedly higher in patients with HCC than in patients with CHB and HCs. Furthermore, the plasma soluble WISP1 in patients with HCC was obviously lower than in that in patients with CHB and HCs. Alpha-fetoprotein (AFP) is a widely recognized biomarker to diagnose HCC which lacks enough sensitivity and specificity. WISP1 promoter methylation status combined with AFP significantly improved the diagnostic ability in discriminating HCC from CHB compared with AFP or WISP1 methylation status alone. In conclusion, hypomethylation of the WISP1 gene promoter may serve as a noninvasive biomarker for detecting HBV-associated HCC.

Plasma diamine oxidase level predicts 6-month readmission for patients with hepatitis B virus-related decompensated cirrhosis.

BACKGROUND AND AIMS: Hepatitis B virus-related decompensated cirrhosis is difficult to cure but has a high readmission rate due to multiple complications. Our aim was to investigate the diagnostic potential value of plasma diamine oxidase (DAO) for 6-month readmission of patients with HBV-related decompensated cirrhosis. METHODS: A total of 135 patients with HBV-related decompensated cirrhosis were prospectively collected at the onset of discharge of hospital, and then were followed up for at least 6 months with the readmission as the primary outcome. The plasma DAO level was measured using enzyme linked immunosorbent assay. In addition, 120 age and sex matched patients with HBV-related compensated cirrhosis were included as controls. RESULTS: A total of 36 patients (36.7%) with decompensated cirrhosis admitted to hospital during the 6-month follow up. The plasma DAO level of readmission group [21.1 (14.5; 29.0) ng/ml] was significantly higher than that in the non-readmission group [12.7 (9.3; 18.0) ng/mL, P < 0.001]. Multivariate analysis showed that the plasma DAO level (HR = 1.102, P < 0.05) and hepatic encephalopathy (HE) (HR = 5.018, P < 0.05) were independent factors for 6-month readmission of decompensated cirrhosis. DAO level showed higher area under the curve of receiver operating characteristic (AUROC) than HE (0.769 vs. 0.598, P < 0.05) and Child-Pugh-Turcotte (CPT) score (0.769 vs. 0.652, P < 0.05) for predicting 6-month readmission rate, with the best cut-off value as 19.7 ng/mL. Furthermore, plasma DAO level (HR = 1.184, P < 0.05) was an independent factor and has the higher AUROC than CPT score for the onset of recurrent HE (0.905 vs. 0.738, P < 0.05) during the 6-month follow up. CONCLUSIONS: Plasma DAO level > 19.7 ng/mL predicts high rate of 6-month readmission in patients with HBV-related decompensated cirrhosis.

Tumor necrosis factor-α-induced protein 8-like 2 mRNA in peripheral blood mononuclear cells is associated with the disease progression of chronic hepatitis B virus infection.

BACKGROUND: Tumor necrosis factor-alpha-induced protein 8-like 2 (TIPE2) is a novel target and molecule in the negative regulation of immune homeostasis. The present study aimed to investigate the dynamic expression of TIPE2 mRNA during the progression of chronic hepatitis B virus (HBV) infection. METHODS: A total of 193 patients with chronic HBV infection were retrospectively recruited into this cross-sectional study, including 97 patients with chronic hepatitis B (CHB), 55 with liver cirrhosis and 41 with HBV-related hepatocellular carcinoma (HCC). TIPE2 mRNA was determined using real-time quantitative polymerase chain reaction. RESULTS: The expression of TIPE2 levels in patients with HCC was significantly decreased compared with expression in patients with liver cirrhosis, CHB and healthy controls (P < 0.05); meanwhile, the TIPE2 mRNA levels in patients with CHB and liver cirrhosis were significantly increased compared with levels in healthy controls (P < 0.01). In liver cirrhosis, the TIPE2 mRNA level in the decompensated state was significantly higher than that in the compensated state (P < 0.05). In HCC patients, TIPE2 mRNA was significantly associated with venous invasion, tumor size and tumor node metastasis stage. Furthermore, the optimal cutoff of 0.78 for the level of TIPE2 mRNA has a sensitivity of 97.56% and a specificity of 88.16% for discriminating HCC from patients with CHB and liver cirrhosis. CONCLUSIONS: TIPE2 mRNA was associated with various stages of chronic HBV infection, ranging from CHB to liver cirrhosis and HCC. Furthermore, TIPE2 mRNA with an optional cutoff value of 0.78 might serve as a promising biomarker to discriminate HBV-associated HCC from CHB and LC patients.