Prevalence and risk factors of covert hepatic encephalopathy in cirrhotic patients: A multicenter study in China.

OBJECTIVE: We aimed to investigate the prevalence of covert hepatic encephalopathy (CHE) in cirrhotic patients in China and its risk factors. METHODS: A multicenter prospective observational study was conducted from January 2021 to March 2022 at 16 medical centers across China to investigate the risk factors of CHE and establish a prediction model for CHE episodes. RESULTS: A total of 528 patients were enrolled in the study. Based on both the psychometric hepatic encephalopathy score and Stroop test results, the prevalence of CHE was 50.4% (266/528), and the consistency between these two tests was 68.9%. Multivariate analysis showed that age (odds ratio [OR] 1.043, 95% confidence interval [CI] 1.022-1.063, P < 0.001), duration of education (OR 0.891, 95% CI 0.832-0.954, P = 0.001), comorbidities of cardiovascular diseases, hypertension, cerebral apoplexy or diabetes mellitus (OR 2.072, 95% CI 1.370-3.133, P < 0.001), Child-Pugh score (OR 1.142, 95% CI 1.029-1.465, P = 0.025), and blood urea nitrogen concentration (OR 1.126, 95% CI 1.038-1.221, P = 0.004) were associated with CHE episodes. According to the Chronic Liver Disease Questionnaire, CHE patients had lower scores for abdominal symptoms and systemic symptoms (P < 0.001), indicating a poor health-related quality of life. Based on a stepwise Cox regression hazard model, we established a nomogram for determining the probabilities of CHE episodes, and the area under the receiver operating characteristic curve was 0.733 (95% CI 0.679-0.788) and 0.713 (95% CI 0.628-0.797) in the training and validation cohorts. CONCLUSIONS: CHE is a common complication of cirrhosis in China. Large-scale studies with long-term follow-up are needed to determine the natural history of Chinese CHE patients.

SHARPIN stabilizes ß-catenin through a linear ubiquitination-independent manner to support gastric tumorigenesis.

BACKGROUND: Aberrant activation of Wnt/ß-catenin signaling by dysregulated post-translational protein modifications, especially ubiquitination is causally linked to cancer development and progression. Although Lys48-linked ubiquitination is known to regulate Wnt/ß-catenin signaling, it remains largely obscure how other types of ubiquitination, such as linear ubiquitination governs its signaling activity. METHODS: The expression and regulatory mechanism of linear ubiquitin chain assembly complex (LUBAC) on Wnt/ß-catenin signaling was examined by immunoprecipitation, western blot and immunohistochemical staining. The ubiquitination status of ß-catenin was detected by ubiquitination assay. The impacts of SHARPIN, a core component of LUBAC on malignant behaviors of gastric cancer cells were determined by various functional assays in vitro and in vivo. RESULTS: Unlike a canonical role in promoting linear ubiquitination, SHARPIN specifically interacts with ß-catenin to maintain its protein stability. Mechanistically, SHARPIN competes with the E3 ubiquitin ligase ß-Trcp1 for ß-catenin binding, thereby decreasing ß-catenin ubiquitination levels to abolish its proteasomal degradation. Importantly, SHARPIN is required for invasiveness and malignant growth of gastric cancer cells in vitro and in vivo, a function that is largely dependent on its binding partner ß-catenin. In line with these findings, elevated expression of SHARPIN in gastric cancer tissues is associated with disease malignancy and correlates with ß-catenin expression levels. CONCLUSIONS: Our findings reveal a novel molecular link connecting linear ubiquitination machinery and Wnt/ß-catenin signaling via SHARPIN-mediated stabilization of ß-catenin. Targeting the linear ubiquitination-independent function of SHARPIN could be exploited to inhibit the hyperactive ß-catenin signaling in a subset of human gastric cancers.

The efficacy of low molecular weight heparin in severe acute pancreatitis: A systematic review and meta-analysis of randomized controlled trials.

OBJECTIVE: The effects of low molecular weight heparin (LMWH) on severe acute pancreatitis (SAP) have been controversial. We aimed to evaluate the efficacy of LMWH on prognosis of SAP by systematic review and meta-analysis. METHODS: We searched relevant studies published up to March 2019 in five databases (MEDLINE/PubMed, EMBASE, the Cochrane Central Register of Controlled Trials in Cochrane Library, China National Knowledge Infrastructure, and the Chinese Journal of Science and Technology of VIP database). RESULTS: Sixteen randomized controlled trials with 1625 patients were included in the final analysis. Most studies were from China. In analysis of laboratory parameters and clinical scores, SAP patients receiving LMWH treatment had lower white blood cell counts, C-reactive protein level, Acute Physiology and Chronic Health Evaluation II score, and computed tomography severity index. In clinical outcomes, SAP patients who received LMWH treatment had shorter hospital stay (pooled mean difference [95% confidence interval; CI] -8.79 [-11.18, -6.40], P < .01), lower mortality (pooled risk ratio [RR] [95% CI] 0.33 [0.24-0.44], P < .01), lower incidences of multiple organ failure (pooled RR [95% CI] 0.34 [0.23-0.52], P < .01), pancreatic pseudocyst (pooled RR [95% CI] 0.49 [0.27-0.90], P = .02), and operation rate (pooled RR [95% CI] 0.39 [0.31-0.50], P < .01). CONCLUSIONS: LMWH could improve the prognosis of SAP, and has a potential role in reducing hospital stay, mortality, incidences of multiple organ failure, pancreatic pseudocyst, and operation rate.

Artificial neural networks accurately predict intra-abdominal infection in moderately severe and severe acute pancreatitis.

OBJECTIVE: The aim of this study was to evaluate the efficacy of artificial neural networks (ANN) in predicting intra-abdominal infection in moderately severe (MASP) and severe acute pancreatitis (SAP) compared with that of a logistic regression model (LRM). METHODS: Patients suffering from MSAP or SAP from July 2014 to June 2017 in three affiliated hospitals of the Army Medical University in Chongqing, China, were enrolled in this study. A univariate analysis was used to determine the different parameters between patients with and without intra-abdominal infection. Subsequently, these parameters were used to build LRM and ANN. RESULTS: Altogether 263 patients with MSAP or SAP were enrolled in this retrospective study. A total of 16 parameters that differed between patients with and without intra-abdominal infection were used to construct both models. The sensitivity of ANN and LRM was 80.99% (95% confidence interval [CI] 72.63-87.33) and 70.25% (95% CI 61.15-78.04), respectively (P > 0.05), whereas the specificity was 89.44% (95% CI 82.89-93.77) and 77.46% (95% CI 69.54-83.87), respectively (P < 0.05). ANN predicted the risk of intra-abdominal infection better than LRM (area under the receiver operating characteristic curve: 0.923 [0.883-0.952] vs 0.802 [0.749-0.849], P < 0.001). CONCLUSIONS: ANN accurately predicted intra-abdominal infection in MSAP and SAP and is an ideal tool for predicting intra-abdominal infection in such patients. Coagulation parameters played an important role in such prediction.

Development and validation of three machine-learning models for predicting multiple organ failure in moderately severe and severe acute pancreatitis.

BACKGROUND: Multiple organ failure (MOF) is a serious complication of moderately severe (MASP) and severe acute pancreatitis (SAP). This study aimed to develop and assess three machine-learning models to predict MOF. METHODS: Patients with MSAP and SAP who were admitted from July 2014 to June 2017 were included. Firstly, parameters with significant differences between patients with MOF and without MOF were screened out by univariate analysis. Then, support vector machine (SVM), logistic regression analysis (LRA) and artificial neural networks (ANN) models were constructed based on these factors, and five-fold cross-validation was used to train each model. RESULTS: A total of 263 patients were enrolled. Univariate analysis screened out sixteen parameters referring to blood volume, inflammatory, coagulation and renal function to construct machine-learning models. The predictive efficiency of the optimal combinations of features by SVM, LRA, and ANN was almost equal (AUC = 0.840, 0.832, and 0.834, respectively), as well as the Acute Physiology and Chronic Health Evaluation II score (AUC = 0.814, P > 0.05). The common important predictive factors were HCT, K-time, IL-6 and creatinine in three models. CONCLUSIONS: Three machine-learning models can be efficient prognostic tools for predicting MOF in MSAP and SAP. ANN is recommended, which only needs four common parameters.

BRD4 Promotes Gastric Cancer Progression and Metastasis through Acetylation-Dependent Stabilization of Snail.

Cancer metastasis, a leading cause of death in patients, is associated with aberrant expression of epigenetic modifiers, yet it remains poorly defined how epigenetic readers drive metastatic growth and whether epigenetic readers are targetable to control metastasis. Here, we report that bromodomain-containing protein 4 (BRD4), a histone acetylation reader and emerging anticancer therapeutic target, promotes progression and metastasis of gastric cancer. The abundance of BRD4 in human gastric cancer tissues correlated with shortened metastasis-free gastric cancer patient survival. Consistently, BRD4 maintained invasiveness of cancer cells in vitro and their dissemination at distal organs in vivo. Surprisingly, BRD4 function in this context was independent of its putative transcriptional targets such as MYC or BCL2, but rather through stabilization of Snail at posttranslational levels. In an acetylation-dependent manner, BRD4 recognized acetylated lysine 146 (K146) and K187 on Snail to prevent Snail recognition by its E3 ubiquitin ligases FBXL14 and ß-Trcp1, thereby inhibiting Snail polyubiquitination and proteasomal degradation. Accordingly, genome-wide transcriptome analyses identified that BRD4 and Snail regulate a partially shared metastatic gene signature in gastric cancer cells. These findings reveal a noncanonical posttranscriptional regulatory function of BRD4 in maintaining cancer growth and dissemination, with immediate translational implications for treating gastric metastatic malignancies with clinically available bromodomain inhibitors. SIGNIFICANCE: These findings reveal a novel posttranscriptional regulatory function of the epigenetic reader BRD4 in cancer metastasis via stabilizing Snail, with immediate translational implication for treating metastatic malignancies with clinically available bromodomain inhibitors. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/79/19/4869/F1.large.jpg.

Norrin maintains malignancy of gastric cancer cells in part through activating AKT signaling.

Human tumorigenesis resembles embryogenesis by aberrant activation of several developmental pathways including Wnt/ß-catenin signaling. Norrin is an atypical ligand for Frizzled receptor that is preferentially expressed in the endothelium to promote retinal vascularization during development. However, its expression pattern and potential roles in human cancers remain unclear. Here we report that Norrin expression is elevated in the parenchymal cells, but not endothelial cells, in gastric cancer (GC). Moreover, Norrin is required for growth and invasion of GC cells and its expression status is associated with unfavorable outcomes. However, analysis of the TGCA database demonstrates that Norrin expression status is not correlated with key target genes of Wnt/ß-catenin signaling. Among several signaling pathways hyperactivated in cancer, Norrin-depleted GC cells also display down-regulated AKT signaling except the canonical Wnt/ß-catenin signaling. Consistently, small molecule-induced cytosolic activation of AKT partially rescues the proliferative and invasive capability of Norrin-depleted cells. Together, these findings suggest a novel role of Norrin in gastric tumorigenesis that could be exploited for adjuvant therapy against the deadly malignancy.

Fecal Microbiota Transplantation Ameliorates Experimentally Induced Colitis in Mice by Upregulating AhR.

Ulcerative colitis (UC) is a chronic non-specific inflammatory disease that occurs in the colon and rectum. While fecal microbiota transplantation (FMT) is gaining attention as a clinical treatment of UC, the molecular mechanisms behind this effect have yet to be fully understood. A C57BL/6 mouse model was established to test whether FMT promotes the recovery of colon inflammation. Administration of 2% dextran sulfate sodium (DSS) for 7 days successfully induced acute colitis, as evidenced by diarrhea, hematochezia and colon shortening as well as a decrease in body weight. FMT alleviated the severity of colon mucosa injury and improved histological alterations compared with that of the DSS group. In addition, FMT promoted homeostasis of the intestinal microbiota. Furthermore, FMT upregulated the expression of aryl hydrocarbon receptor (AHR), interleukin-10 (IL-10), and transforming growth factor beta (TGF-ß) in colon tissues. These results suggest that the significant anti-inflammatory effect of FMT may be attributed to its promotion of IL-10 and TGF-ß production and AHR activation. Based on these results, FMT had a favorable therapeutic effect on DSS-induced colitis.

SHP-1 Acts as a Tumor Suppressor in Hepatocarcinogenesis and HCC Progression.

Src homology region 2 (SH2) domain-containing phosphatase 1 (SHP-1, also known as PTPN6) is a nonreceptor protein tyrosine phosphatase that acts as a negative regulator of inflammation. Emerging evidence indicates that SHP-1 plays a role in inhibiting the progression of hepatocellular carcinoma (HCC). However, the role of SHP-1 in hepatocarcinogenesis remains unknown. Here, we find that levels of SHP-1 are significantly downregulated in human HCC tissues compared with those in noncancerous tissues (P < 0.001) and inversely correlate with tumor diameters (r = -0.4130, P = 0.0002) and serum α-fetoprotein levels (P = 0.047). Reduced SHP-1 expression was associated with shorter overall survival of patients with HCC with HBV infection. Overexpression of SHP-1 suppressed proliferation, migration, invasion, and tumorigenicity of HCC cells, whereas knockdown of SHP-1 enhanced the malignant phenotype. Moreover, knockout of Ptpn6 in hepatocytes (Ptpn6HKO ) enhanced hepatocarcinogenesis induced by diethylnitrosamine (DEN) as well as metastasis of primary liver cancer in mice. Furthermore, systemic delivery of SHP-1 by an adenovirus expression vector exerted a therapeutic effect in an orthotopic model of HCC in NOD/SCID mice and DEN-induced primary liver cancers in Ptpn6HKO mice. In addition, SHP-1 inhibited the activation of JAK/STAT, NF-κB, and AKT signaling pathways, but not the MAPK pathway in primary hepatocytes from DEN-treated mice and human HCC cells. Together, our data implicate SHP-1 as a tumor suppressor of hepatocarcinogenesis and HCC progression and propose it as a novel prognostic biomarker and therapeutic target of HCC.Significance: The nonreceptor protein tyrosine phosphatase SHP-1 acts as a tumor suppressor in hepatocellular carcinoma. Cancer Res; 78(16); 4680-91. ©2018 AACR.

Epigenetic restriction of Hippo signaling by MORC2 underlies stemness of hepatocellular carcinoma cells.

The evolutionarily conserved Hippo signaling pathway is a key regulator of stem cell self-renewal, differentiation, and organ size. While alterations in Hippo signaling are causally linked to uncontrolled cell growth and a broad range of malignancies, genetic mutations in the Hippo pathway are uncommon and it is unclear how the tumor suppressor function of the Hippo pathway is disrupted in human cancers. Here, we report a novel epigenetic mechanism of Hippo inactivation in the context of hepatocellular carcinoma (HCC). We identify a member of the microrchidia (MORC) protein family, MORC2, as an inhibitor of the Hippo pathway by controlling upstream Hippo regulators, neurofibromatosis 2 (NF2) and kidney and brain protein (KIBRA). Mechanistically, MORC2 forms a complex with DNA methyltransferase 3A (DNMT3A) at the promoters of NF2 and KIBRA, leading to their DNA hyper-methylation and transcriptional repression. As a result, NF2 and KIBRA are crucial targets of MORC2 to regulate confluence-induced activation of Hippo signaling and contact inhibition of cell growth under both physiological and pathological conditions. The MORC2-NF2/KIBRA axis is critical for maintaining self-renewal, sorafenib resistance, and oncogenicity of HCC cells in vitro and in nude mice. Furthermore, MORC2 expression is elevated in HCC tissues, associated with stem-like properties of cancer cells, and disease progression in patients. Collectively, MORC2 promotes cancer stemness and tumorigenesis by facilitating DNA methylation-dependent silencing of Hippo signaling and could be a potential molecular target for cancer therapeutics.

The HNF1α-regulated lncRNA HNF1A-AS1 reverses the malignancy of hepatocellular carcinoma by enhancing the phosphatase activity of SHP-1.

BACKGROUND: Our previous study has demonstrated that hepatocyte nuclear factor 1α (HNF1α) exerts potent therapeutic effects on hepatocellular carcinoma (HCC). However, the molecular mechanisms by which HNF1α reverses HCC malignancy need to be further elucidated. METHODS: lncRNA microarray was performed to identify the long noncoding RNAs (lncRNAs) regulated by HNF1α. Chromatin immunoprecipitation and luciferase reporter assays were applied to clarify the mechanism of the transcriptional regulation of HNF1α to HNF1A antisense RNA 1 (HNF1A-AS1). The effect of HNF1A-AS1 on HCC malignancy was evaluated in vitro and in vivo. RNA pulldown, RNA-binding protein immunoprecipitation and the Bio-Layer Interferometry assay were used to validate the interaction of HNF1A-AS1 and Src homology region 2 domain-containing phosphatase 1 (SHP-1). RESULTS: HNF1α regulated the expression of a subset of lncRNAs in HCC cells. Among these lncRNAs, the expression levels of HNF1A-AS1 were notably correlated with HNF1α levels in HCC cells and human HCC tissues. HNF1α activated the transcription of HNF1A-AS1 by directly binding to its promoter region. HNF1A-AS1 inhibited the growth and the metastasis of HCC cells in vitro and in vivo. Moreover, knockdown of HNF1A-AS1 reversed the suppressive effects of HNF1α on the migration and invasion of HCC cells. Importantly, HNF1A-AS1 directly bound to the C-terminal of SHP-1 with a high binding affinity (KD = 59.57 ± 14.29 nM) and increased the phosphatase activity of SHP-1. Inhibition of SHP-1 enzymatic activity substantially reversed the HNF1α- or HNF1A-AS1-induced reduction on the metastatic property of HCC cells. CONCLUSIONS: Our data revealed that HNF1A-AS1 is a direct transactivation target of HNF1α in HCC cells and involved in the anti-HCC effect of HNF1α. HNF1A-AS1 functions as phosphatase activator through the direct interaction with SHP-1. These findings suggest that regulation of the HNF1α/HNF1A-AS1/SHP-1 axis may have beneficial effects in the treatment of HCC.

Apoptosis signal-regulating kinase 1 mediates the inhibitory effect of hepatocyte nuclear factor-4α on hepatocellular carcinoma.

Previous studies provided substantial evidence of a striking suppressive effect of hepatocyte nuclear factor 4α (HNF4α) on hepatocellular carcinoma (HCC). Apoptosis signal-regulating kinase 1 (ASK1) is involved in death receptor-mediated apoptosis and may acts as a tumor suppressor in hepatocarcinogenesis. However, the status and function of ASK1 during HCC progression are unclear. In this study, we found that HNF4α increased ASK1 expression by directly binding to its promoter. ASK1 expression was dramatically suppressed and correlated with HNF4α levels in HCC tissues. Reduced ASK1 expression was associated with aggressive tumors and poor prognosis for human HCC. Moreover, ASK1 inhibited the malignant phenotype of HCC cells in vitro. Intratumoral ASK1 injection significantly suppressed the growth of subcutaneous HCC xenografts in nude mice. More interestingly, systemic ASK1 delivery strikingly inhibited the growth of orthotopic HCC nodules in NOD/SCID mice. In addition, inhibition of endogenous ASK1 partially reversed the suppressive effects of HNF4α on HCC. Collectively, this study highlights the suppressive effect of ASK1 on HCC and its biological significance in HCC development. These outcomes broaden the knowledge of ASK1 function in HCC progression, and provide a novel potential prognostic biomarker and therapeutic target for advanced HCC.