Background: G-protein-coupled bile acid receptor Gpbar1 (TGR5) is a newly identified liver tumor suppressor in carcinogenesis. This present study was therefore to determine the potential value of serum TGR5 promoter methylation in identifying hepatocellular carcinoma (HCC) from chronic hepatitis B (CHB) patients.Methods: The circulating cell-free DNA (cfDNA) was extracted from a retrospective dataset including 160 HCC, 88 CHB and 45 healthy controls (HCs). Methylation status of TGR5 promoter was examined by methylation-specific polymerase chain reaction (MSP).Results: Hypermethylation of the TGR5 promoter occurred significantly more frequent in HCC (77/160, 48.13%) than CHB (12/88, 13.64%; p<0.01) and HCs (2/45, 4.44%; p<0.01). The methylation rate of TGR5 in HCC patients ≥60 years old was significantly higher than those <60 years old (p<0.05). Alpha fetoprotein (AFP) had sensitivity of 58.13%, 30.63% and 24.38% at cut-off points of 20, 200 and 400ng/ml respectively; while TGR5 methylation combined AFP had sensitivity of 81.25%, 68.13% and 65%. AFP had specificity of 47.73%, 92.05% and 98.86% at cut-off points of 20, 200 and 400ng/ml respectively; while TGR5 methylation combined AFP had specificity of 38.64%, 78.41% and 85.23%. AFP had Youden index of 0.06, 0.23 and 0.23 at cut-off points of 20, 200 and 400ng/ml respectively; while TGR5 methylation combined AFP had Youden index of 0.20, 0.47 and 0.50.Conclusions: Our findings strongly suggested the combination of serum TGR5 promoter methylation and AFP enhanced the diagnostic value of AFP alone in discriminating HCC from CHB patients.

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The role of epidermal growth factor-containing fibulin-like extracellular matrix protein 1 (EFEMP1) inhibiting migration in hepatocellular carcinoma (HCC) remains unknown. Expression of EFEMP1 in HCC cell lines were quantified by western blotting and real-time PCR. The role of EFEMP1 in HCC cell migration was explored in vitro via siRNA and adding purified EFEMP1 protein. The associated molecule expression was detected by western blotting after downregulation of EFEMP1 and also tested by immunohistochemistry. Eight pairs of HCC non-HCC liver samples and 215 HCC samples were subjected to immunohistochemistry. EFEMP1 was highly expressed in 7,721 and HepG2 HCC cell lines while HuH7 HCC cell line expressed the lowest level of EFEMP1 compared with the others. Downregulating EFEMP1 by siRNA markedly increased the migration ability of HCC cells while adding purified EFEMP1 protein inhibited HCC cell migration. Downregulation of EFEMP1 increased the expression of ERK1/2, MMP2 and MMP9. Furthermore, U0126 (a highly selective and potent inhibitor of pERK1/2) could abrogate the migration ability enhanced by siRNA. Accordingly, MMP2 and MMP9 were inversely expressed with EFEMP1 expression by immunohistochemistry. EFEMP1 downregulated in HCC tissues, and lower EFEMP1 expression was significantly associated with HCC patients with ascites (P=0.050), vascular invasion (P=0.044), poorer differentiation (P=0.002) and higher clinical stage (P=0.003).

Aberrant sera DNA methylation of CDH1, DNMT3b and ESR1 gene promoters could be a biomarker in the early diagnosis of HBV-related HCC.

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related mortality worldwide. Epigenetic analysis has attracted increasing attention in the molecular diagnosis of HCC. Cysteine dioxygenase 1 (CDO1) is a key enzyme in the taurine biosynthetic pathway and converts cysteine to cysteine sulfinate. The CDO1 gene is a tumor suppressor gene and is usually silenced by the methylation of its promoter in carcinogenesis. In this study, we evaluated whether the methylation status of CDO1 gene promoter is of diagnostic value for hepatitis B virus (HBV)-related HCC. The CDO1 promoter methylation status was determined in serum samples using methylation-specific polymerase chain reaction (MSP) in a cohort of 123 patients with HBV-related HCC, 28 with liver cirrhosis (LC), 29 with chronic hepatitis B (CHB) and 20 healthy controls. The frequency of the CDO1 promoter methylation in HBV-related HCC (42.3%) was significantly higher than that in LC (14.3%), CHB (6.9%) and healthy controls (0%) (P = 0.006; P < 0.0001; P < 0.0001; respectively). Furthermore, in HCC patients, the frequency of CDO1 promoter methylation was higher in advanced stages (III-IV) (53%) than the early stages (I-II) (20%) (P = 0.001). Evaluation of the CDO1 promoter methylation status in serum, in combination with AFP (> 20 ng/ml), significantly improved the diagnostic value, with sensitivity and specificity of 82.9% and 75.4%, respectively in distinguishing HCC from LC and CHB. In conclusion, methylation status of serum CDO1 gene promoter may be helpful in the diagnosis of HCC and the estimation of the HCC stages.

Coronavirus disease 2019 (COVID-19) is considered to be spread primarily by people who have tested positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we discuss a patient with severe COVID-19 and a history of type 2 diabetes who had a recurrence of positive SARS-CoV-2 ribonucleic acid (RNA) after recovering. The patient was initially discharged after two consecutive negative SARS-CoV-2 RNA tests and partially absorbed bilateral lesions on chest computed tomography (CT). However, at his first follow-up, reverse transcription-polymerase chain reaction (RT-PCR) assay with an oropharyngeal swab sample was positive for SARS-CoV-2. Despite this, he displayed no obvious clinical symptoms and improved chest CT. The patient was prescribed anti-viral medication. Eight consecutive RT-PCR assays on oropharyngeal swab specimens were conducted after he was re-admitted to our hospital. The results tested positive on the 12th, 14th, 19th, 23rd and 26th of March and negative on the 28th of March, and 6th and 12th of April. After his second discharge, he has tested negative for 5 weeks. This case highlights the importance of active surveillance of SARS-CoV-2 RNA during the follow-up period so that an infectivity assessment can be made.

Tumor endothelial marker 8 (TEM8) was recently suggested as a putative anti-tumor target in several types of human cancer based on its selective overexpression in tumor versus normal endothelial cells. The objective of this study was to detect the potential functions of TEM8 in osteosarcoma. Overall, TEM8 was mainly located in cytoplasm and was up-regulated in osteosarcoma compared to benign bone lesions and adjacent non tumor tissue (ANT). High TEM8 expression group had a significant lower overall survival rate than that in the low TEM8 expression group. TEM8 knock-down by siRNA or shRNA results in significant reduction of osteosarcoma cell growth and proliferation both in vitro and in vivo. Ablation of TEM8 led to increasing of p21 and p27 and suppression of cyclin D1 mediated by Erk1/2 activity. These findings suggest that down-regulation of TEM8 play an important role in the inhibition of tumorigenesis and development of osteosarcoma.

Suppressor of cytokine signaling (SOCS) 1 plays a crucial role in the immune response and might contribute to the prognoses of liver failure treated with glucocorticoid. We recruited 47 acute-on-chronic hepatitis B liver failure (ACHBLF) patients receiving glucocorticoid treatment and 30 healthy controls to determine the potential effects of glucocorticoid on the transcriptional level of SOCS1 in peripheral blood mononuclear cells. On the third and twenty-eighth days of glucocorticoid treatment, SOCS1 expression was negatively correlated with model for end-stage liver disease (MELD) score. Interleukin-6 (IL-6) and tumor-necrosis factor-a (TNF-a) levels were statistically lower, while the SOCS1 transcription level was higher in survivors than non-survivors both in pre-and post-treatment ACHBLF patients. The methylation rate of the SOCS1 promoter in ACHBLF patients was higher than in healthy control patients as determined by methylation-specific polymerase chain reaction. The mRNA level of SOCS1 in methylated promoters was significantly lower than from patients with unmethylated SOCS1 promoters. interferon (IFN)-c-responsive and STAT1-dependent gene expression was higher in survivors and was dramatically decreased with rising expression of SOCS1 after glucocorticoid treatment. Mortality rates were significantly higher in methylated patients than for those without methylation at the end of a 90-day follow-up. Furthermore, we found that five in six surviving patients displayed demethylated SOCS1 on the twenty-eighth day after treatment, while that number was 3 in 10 in the non-survivors. These findings suggested that ACHBLF patients without SOCS1 methylation may have a favorable response to corticosteroid treatment.