FXR Maintains the Intestinal Barrier and Stemness by Regulating CYP11A1-Mediated Corticosterone Synthesis in Biliary Obstruction Diseases.

Biliary obstruction diseases are often complicated by an impaired intestinal barrier, which aggravates liver injury. Treatment of the intestinal barrier is often neglected. To investigate the mechanism by which intestinal bile acid deficiency mediates intestinal barrier dysfunction after biliary obstruction and identify a potential therapeutic modality, we mainly used a bile duct ligation (BDL) mouse model to simulate biliary obstruction and determine the important role of the bile acid receptor FXR in maintaining intestinal barrier function and stemness. Through RNA-seq analysis of BDL and sham mouse crypts and qRT-PCR performed on intestinal epithelial-specific Fxr knockout (FxrΔIEC) and wild-type mouse crypts, we found that FXR might maintain intestinal stemness by regulating CYP11A1 expression. Given the key role of CYP11A1 during glucocorticoid production, we also found that FXR activation could promote intestinal corticosterone (CORT) synthesis by ELISA. Intestinal organoid culture showed that an FXR agonist or corticosterone increased crypt formation and organoid growth. Further animal experiments showed that corticosterone gavage treatment could maintain intestinal barrier function and stemness, decrease LPS translocation, and attenuate liver injury in BDL mice. Our study hopefully provides a new theoretical basis for the prevention of intestinal complications and alleviation of liver injury after biliary obstruction.

Targeting anillin inhibits tumorigenesis and tumor growth in hepatocellular carcinoma via impairing cytokinesis fidelity.

Targeting cytokinesis can suppress tumor growth by blocking cell division and promoting apoptosis. We aimed to characterize key cytokinesis regulator in hepatocellular carcinoma (HCC) progression, providing insights into identifying promising HCC therapeutic targets. The unbiased bioinformatic screening identified Anillin actin binding protein (ANLN) as a critical cytokinesis regulator involved in HCC development. Functional assay demonstrated that knockdown of ANLN inhibited HCC growth by inducing cytokinesis failure and DNA damage, leading to multinucleation and mitotic catastrophe. Mechanistically, ANLN acts as a scaffold to strengthen interaction between RACGAP1 and PLK1. ANLN promotes PLK1-mediated RACGAP1 phosphorylation and RhoA activation to ensure cytokinesis fidelity. To explore the function of ANLN in HCC tumorigenesis, we hydrodynamically transfected c-Myc and NRAS plasmids into Anln+/+, Anln+/-, and Anln-/- mice through tail vein injection. Hepatic Anln ablation significantly impaired c-Myc/NRAS-driven hepatocarcinogenesis. Moreover, enhanced hepatic polyploidization was observed in Anln ablation mice, manifesting as increasing proportion of cellular and nuclear polyploidy. Clinically, ANLN is upregulated in human HCC tissues and high level of ANLN is correlated with poor patients' prognosis. Additionally, the proportion of cellular polyploidy decreases during HCC progression and ANLN level is significantly correlated with cellular polyploidy proportion in human HCC samples. In conclusion, ANLN is identified as a key cytokinesis regulator contributing to HCC initiation and progression. Our findings revealed a novel mechanism of ANLN in the regulation of cytokinesis to promote HCC tumorigenesis and growth, suggesting targeting ANLN to inhibit cytokinesis may be a promising therapeutic strategy for HCC.

Hypermethylation of GNA14 and its tumor-suppressive role in hepatitis B virus-related hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most lethal cancers worldwide, and its specific mechanism has not been fully elucidated. Inactivation of tumor suppressors may contribute to the occurrence, progression, and recurrence of HCC. DNA methylation is a crucial mechanism involved in regulating the occurrence of HCC. Herein, we aimed to identify the key methylation-related tumor suppressors as well as potential biomarkers and therapeutic targets in HCC. Methods: Combined analysis of TCGA and GEO databases was performed to obtain potential methylation-related tumor suppressors in HCC. Methyl-target sequencing was performed to analyze the methylation level of the GNA14 promoter. The diagnostic value of GNA14 as a predictor of HCC was evaluated in HCC tumor samples and compared with normal tissues. The functional role of GNA14 and its upstream and downstream regulatory factors were investigated by gain-of-function and loss-of-function assays in vitro. Subcutaneous tumorigenesis, lung colonization, and orthotopic liver tumor model were performed to analyze the role of GNA14 in vivo.Results: The expression of GNA14 was found to be downregulated in HCC and it was negatively correlated with hepatitis B virus (HBV) infection, vascular invasion, and prognosis of HCC. DNA methylation was demonstrated to be responsible for the altered expression of GNA14 and was regulated by HBV-encoded X protein (HBx). GNA14 regulated the RB pathway by promoting Notch1 cleavage to inhibit tumor proliferation, and might inhibit tumor metastasis by inhibiting the expression of JMJD6. Conclusion: GNA14 could be regulated by HBx by modulating the methylation status of its promoter. We identified GNA14 as a potential biomarker and therapeutic target for HCC.

EAG1 enhances hepatocellular carcinoma proliferation by modulating SKP2 and metastasis through pseudopod formation.

Ether-à-go-go-1 (EAG1), one of the potassium channels, is involved in various physiological processes and plays an important role in the tumorigenesis of many kinds of cancer. EAG1 is highly expressed in hepatocarcinoma cells and is closely related to clinical prognosis, but the molecular mechanism remains elusive. In this study, we verified that EAG1 promotes the proliferation of hepatocellular carcinoma (HCC) both in vitro and in vivo. It promotes cell cycle progression by inhibiting the ubiquitination of SKP2. In addition, EAG1 promotes the migration and invasion of HCC by promoting cell pseudopod formation. Furthermore, in a high-pressure plasmid-injected mouse liver orthotopic carcinoma model, astemizole, an EAG family blocker, can significantly inhibit the formation of liver cancer. Meanwhile, liver-specific EAG1 knockout mice show resistance to hepatocarcinogenesis. This research demonstrated that EAG1 plays an important role in the progression of HCC, and could be a potential therapeutic target for HCC.

A novel role for farnesoid X receptor in the bile acid-mediated intestinal glucose homeostasis.

The farnesoid X receptor (FXR), as a bile acid (BA) sensor, plays an important role in the regulation of lipid metabolism. However, the effects and underlying molecular mechanisms of FXR on intestinal glucose homeostasis remain elusive. Herein, we demonstrated that FXR and glucose transporter 2 (GLUT2) are essential for BA-mediated glucose homeostasis in the intestine. BA-activated FXR enhanced glucose uptake in intestinal epithelial cells by increasing the expression of GLUT2, which depended on ERK1/2 phosphorylation via S1PR2. However, it also reduced the cell energy generation via inhibition of oxidative phosphorylation, which is crucial for intestinal glucose transport. Moreover, BA-activated FXR signalling potently inhibited specific glucose flux through the intestinal epithelium to the circulation, which reduced the increase in blood glucose levels in mice following oral glucose administration. This trend was supported by the changed ratio of GLUT2 to SGLT1 in the brush border membrane (BBM), including especially decreased GLUT2 abundance in the BBM. Furthermore, impaired intestinal FXR signalling was observed in the patients with intestinal bile acid deficiency (IBAD). These findings uncover a novel function by which FXR sustains the intestinal glucose homeostasis and provide a rationale for FXR agonists in the treatment of IBAD-related hyperglycaemia.

Splenic marginal zone lymphoma: a case report and literature review.

BACKGROUND: Splenic marginal zone lymphoma (SMZL) is a rare non-Hodgkin lymphoma, and much little is known about its clinical characteristics and management strategies. Here we present a case of SMZL and review relevant literature to provide a better recognition of this disease entity. CASE PRESENTATION: A 49-year-old Chinese female was admitted to our hospital with complaints of abdominal distension and acid reflux. Physical examinations and imaging investigations suggested the presence of splenomegaly. Laboratory workups revealed mildly reduced white blood cell count otherwise was not remarkable. The patient underwent splenectomy. Histological examination combined with immunohistochemical analysis of the resected spleen confirmed the diagnosis of SMZL. The patient recovered uneventfully during follow-ups. CONCLUSIONS: Due to the rarity and unspecific clinical features, SMZL is extremely challenging to be diagnosed preoperatively. Patients with SMZL are generally associated with favorable prognosis. Combining the staging characteristics of non-Hodgkin's lymphoma and splenic primary lymphoma may assist in clinical staging management of SMZL.

How DNA methylation affects the Warburg effect.

Significant enhancement of the glycolysis pathway is a major feature of tumor cells, even in the presence of abundant oxygen; this enhancement is known as the Warburg effect, and also called aerobic glycolysis. The Warburg effect was discovered nearly a hundred years ago, but its specific mechanism remains difficult to explain. DNA methylation is considered to be a potential trigger for the Warburg effect, as the two processes have many overlapping links during tumorigenesis. Based on a widely recognized potential mechanism of the Warburg effect, we here summarized the relationship between DNA methylation and the Warburg effect with regard to cellular energy metabolism factors, such as glycolysis related enzymes, mitochondrial function, glycolysis bypass pathways, the tumor oxygen sensing pathway and abnormal methylation conditions. We believe that clarifying the relationship between these different mechanisms may further help us understand how DNA methylation works on tumorigenesis and provide new opportunities for cancer therapy.

Inhibiting the NF-κB pathway enhances the antitumor effect of cabazitaxel by downregulating Bcl-2 in pancreatic cancer.

Optimizing the currently available treatment options for pancreatic cancer (PC) is a priority. Cabazitaxel (CTX), a semisynthetic taxane, is mainly used for treating patients with PC who are resistant to paclitaxel (PTX) or docetaxel, due its poor affinity for P­glycoprotein. However, there are only a few studies demonstrating the effect of CTX on PC. The present study aimed to investigate the efficiency and underlying mechanism of CTX in PC treatment. Cell proliferation, colony formation assay and apoptosis analysis were achieved in the two human PC cell lines AsPC­1 and BxPC­3. Drug sensitivity test was performed in BxPC­3 tumor­bearing mice. The results demonstrated that CTX had a lower half maximal inhibitory concentration compared with PTX for the inhibition of cell proliferation, both in vivo and in vitro. Furthermore, the nuclear factor­κB (NF­κB) pathway was activated following cell treatment with CTX, and NF­κB p65 overexpression attenuated CTX cytotoxicity. In addition, the combined use of the specific NF­κB inhibitor caffeic acid phenethyl ester (CAPE) with CTX significantly enhanced CTX effect, both in vivo and in vitro. Similarly, the mRNA and protein expression of B­cell lymphoma-2 was decreased in AsPC­1 and BxPC­3 cells following treatment with CTX and CAPE, suggesting that NF­κB may serve a crucial role in CTX efficiency. In conclusion, results from our previous study indicated that CTX could potentially replace PTX in the treatment of PC, and the present study demonstrated that CTX combination with an NF­κB inhibitor may be considered as a potential therapeutic option for PC, which may improve the prognosis of patients with PC.

EPS8L3 promotes hepatocellular carcinoma proliferation and metastasis by modulating EGFR dimerization and internalization.

As a member of epidermal growth factor receptor (EGFR) kinase substrate 8 (EPS8) family, the role of EPS8 like 3 protein (EPS8L3) has not been well studied in malignancies. However, EPS8 has been reported to be associated with prognosis and functions in several kinds of cancers. Hence, whether EPS8L3 plays similar roles in the tumorigenesis of human cancers, especially in hepatocellular carcinoma (HCC), is still needed to be further explored. In this study, we revealed that EPS8L3 was overexpressed in HCC tissues compared with adjacent non-tumor tissues, and was associated with a poor clinical prognosis. Both in vitro and in vivo experiments showed that EPS8L3 could promote the proliferative ability by downregulating p21/p27 expression, and promote the migratory and invasive abilities by upregulating matrix metalloproteinase-2 expression. Furthermore, we demonstrated that EPS8L3 could affect the activation of the EGFR-ERK pathway by modulating EGFR dimerization and internalization, which may not depend on the formation of EPS8L3-SOS1-ABI1 complex. Taken together, our study showed that EPS8L3 plays a pivotal role in the tumorigenesis and progression of HCC, and it might be a potential therapeutic target for HCC.

Downregulation of MGMT promotes proliferation of intrahepatic cholangiocarcinoma by regulating p21.

BACKGROUND: Intrahepatic cholangiocarcinoma (ICC) is one of the most devastating cancers of the gastrointestinal tract. It is crucial to determine the accurate prognostic factors and find new therapeutic strategies. Meanwhile, O6-methylguanine-DNA methyltransferase (MGMT) is associated with malignant tumor progression. Thus, further studies are needed to investigate whether MGMT plays a similar role in ICC. MATERIALS AND METHODS: Quantitative real-time PCR, western blot, and immunohistochemistry staining were used to detect the expression of MGMT in ICC tissues. The correlations between MGMT expression and clinicopathologic features were analyzed. The cell-proliferation assay and colony-formation assay were applied to evaluate proliferation ability, while methylation-specific PCR were used to detect the methylation status of the MGMT promoter CpG island in ICC tissues and cells. RESULTS: Our study found that the expression of MGMT was decreased in ICC tissues when compared with paired normal tissues. In addition, we demonstrated that MGMT expression was positively correlated with overall survival rates and tumor histological grade. Silencing of MGMT significantly promoted cell proliferation in ICC. Further research showed that silencing of MGMT induced cells to enter S phase by inhibiting p21, p27, and Cyclin E expression, ultimately promoting ICC proliferation. We also demonstrated that the MGMT promoter was highly methylated in ICC, and the levels of MGMT and p21 mRNA increased after DNA demethylation. In addition, the levels of MGMT and p21 protein were positively correlated in ICC tissues. CONCLUSION: MGMT may play a critical role in carcinogenesis and the development of ICC, and provides a new marker of clinical prognosis and target for ICC treatment.

CDCA5, Transcribed by E2F1, Promotes Oncogenesis by Enhancing Cell Proliferation and Inhibiting Apoptosis via the AKT Pathway in Hepatocellular Carcinoma.

Cell division cycle associated 5 (CDCA5) is an important element for the interaction between cohesin and chromatin in interphase. It is abnormally expressed in many types of cancer and works as an indicator of poor prognosis, but little is known about its activity in hepatocellular carcinoma (HCC). In the present study, we found that the expression of CDCA5 was upregulated in HCC tissues compared to paracancerous tissues and had a negative correlation with patient survival. Cell proliferation and tumorigenesis were inhibited and cell apoptosis was induced with the knockdown of CDCA5, suggesting an oncogenic role of CDCA5 in liver cancer. Luciferase reporter assay and chromatin immunoprecipitation showed that CDCA5 was transcribed by E2F1. Furthermore, we confirmed that CDCA5 interrupted cell behavior via the AKT pathway. These findings demonstrated that CDCA5 plays an important role in HCC progression.

The potassium channel KCa3.1 promotes cell proliferation by activating SKP2 and metastasis through the EMT pathway in hepatocellular carcinoma.

The intermediate conductance calcium-activated potassium channel (KCa3.1) plays an important role in maintaining intracellular calcium homeostasis and is involved in the tumorigenesis of many human cancers. However, it is unknown whether KCa3.1 plays a role in the genesis of hepatocellular carcinoma (HCC), one of the most common malignant tumors worldwide with a very poor prognosis. In our study, we found that the expression of KCa3.1 was significantly elevated in poorly differentiated HCC tissues compared to adjacent noncancerous tissues. In vitro and in vivo experiments showed that KCa3.1 could promote cell proliferation, migration, and invasion of HCC. Mechanistically, KCa3.1 promoted cell cycle progression and migration and invasion of HCC cells by activating S-phase protein kinase 2 (SKP2) to trigger the degradation of p21 and p27 and targeting Reelin (RELN) to induce epithelial-mesenchymal transition (EMT), respectively. Taken together, our results demonstrate that KCa3.1 plays an important role in the genesis and progression of HCC, implying that it might be a promising therapeutic target in HCC.

Management of intrahepatic splenosis:a case report and review of the literature.

BACKGROUND: Splenosis is the heterotopic autotransplantation and implantation of splenic tissue after splenic trauma or splenectomy. Considering that splenosis often occurs in the mesentery, omentum, and peritoneum, intrahepatic splenosis has seldom been reported. We report a rare case of isolated intrahepatic splenosis in a 54-year-old man who presented with a liver mass thought to be hepatocellular carcinoma. CASE PRESENTATION: A 54-year-old man was referred to our hospital for further evaluation of a liver lesion. The patient was asymptomatic and had a history of emergent splenectomy after a high-altitude falling accident. Abdominal contrast-enhanced computed tomography revealed a 4.5 × 3.3 cm lesion that was located in segment IV of the left liver lobe. The lesion had an inhomogeneous enhancement during the arterial phase and diminished enhancement during the portal and equilibrium phases. Similar radiological features were also observed on a contrast magnetic resonance imaging scan. Partial hepatectomy was performed with the suspicion of hepatocellular carcinoma. Pathological examination of the liver specimen revealed intrahepatic splenosis. CONCLUSION: Splenosis should be considered in differential diagnosis of a liver mass discovered years after splenic trauma or surgery. A proposed scoring system may be helpful in evaluating the suspicious degree of intrahepatic mass to be splenosis. Invasive treatments are not recommended for asymptomatic patients, since the splenosis can provide beneficial immunologic function.

Inhibition of KLHL21 prevents cholangiocarcinoma progression through regulating cell proliferation and motility, arresting cell cycle and reducing Erk activation.

Kelch-like family member 21 (KLHL21) is involved in cell mitosis and motility. Nevertheless, the clinical significance and biological function of KLHL21 in cholangiocarcinoma (CCA) are elusive. This is the first study to describe a pivotal role for KLHL21 in the progression of CCA. The expression of KLHL21 was elevated in CCA tissues compared with paired normal bile duct tissues. In addition, immunohistochemical and statistical analyses demonstrated that the expression of KLHL21 correlated inversely with tumor histological grade (p < 0.05) and the overall survival of patients (p < 0.01). In CCA cells, we found that the inhibition of KLHL21 significantly reduced proliferation, migration and invasion. Further results indicated that inhibition of KLHL21 triggered G0/G1 cell cycle arrest, leading to the increased expression of P21 and P27 and decreased expression of Cyclin E1, which eventually resulted in proliferation suppression in CCA cells. Furthermore, KLHL21 knockdown alleviated the activation of the Erk signaling pathway via decreasing the expression of phospho-Erk1/2. Our data demonstrated that KLHL21 plays an essential role in the tumorigenesis and progression of CCA, implying that it might serve as a potential therapeutic target for CCA treatment.

KCa3.1 as an Effective Target for Inhibition of Growth and Progression of Intrahepatic Cholangiocarcinoma.

Background: Intrahepatic cholangiocarcinoma (ICC) is a high malignant tumor arising from the bile ducts in the liver with a poor prognosis. As current molecular targeted therapies and systemic chemotherapies had limited success in ICC, novel therapeutic targets are needed. In this study, we attempted to investigate the expression and the role of the intermediate conductance calcium-activated potassium channel (KCa3.1) in ICC. Methods: The expression levels of KCa3.1 channel were measured in 81 resected ICC tumor specimens and the clinicopathological significance of these levels were determined. KCa3.1 channel inhibitor and siRNA were used to study the role of KCa3.1 in proliferation, migration, and invasion of ICC cell lines. The effect of KCa3.1 channel blockade on tumor growth in vivo was also studied using xenograft model in nude mice. Results: The protein expression of KCa3.1 channel was upregulated in ICC tissues and was correlated with age, lymph node metastasis and TNM stage. And high KCa3.1 expression indicated a worse prognosis in ICC patients. Blocking KCa3.1 channel with a specific inhibitor TRAM-34 reduced the proliferation and invasion of ICC cells. Knockdown of KCa3.1 could achieve the same effects through decreasing NF-κB activation. Further in vivo studies demonstrated that KCa3.1 channel blockade suppressed ICC tumor growth. Conclusions: Our observations suggested KCa3.1 might be a promising novel therapeutic target in intrahepatic cholangiocarcinoma.

Bile deficiency induces changes in intestinal glucose absorption in mice.

BACKGROUND: Biliary tract obstruction is a common clinical problem. In this study, we attempted to understand the change in intestinal glucose absorption after biliary tract obstruction. METHODS: Experimental models of murine biliary duct ligation and external biliary drainage were established. Murine intestinal mucosal glucose absorption was examined with Ussing chambers according to the increase in the short-circuit current in vitro and blood glucose measurement after oral glucose in vivo. The protein expression of the sodium-glucose cotransporter (SGLT1) and the facilitated glucose transporter, member 2 (GLUT2) was analyzed by Western blot and immunohistochemistry. RESULTS: The results from Ussing chamber experiments showed that duodenal mucosal glucose absorption levels were significantly higher in biliary duct ligation and biliary drainage mice than those in normal control mice at 1 and 2 weeks after the operation. Gastrointestinal bile acid administration almost reversed the elevated duodenal mucosal glucose absorption to the normal level in biliary drainage mice. The results from the experiments in vivo further confirmed that the glucose absorption increased in biliary duct ligation and biliary drainage mice. The protein expression levels of SGLT1 in the duodenal mucosae of both biliary duct ligation and biliary drainage mice were markedly higher than those in control mice, and the protein expression of GLUT2 was not significantly altered, compared with control mice. CONCLUSION: Bile deficiency in the intestine upregulates the expression of intestinal mucosal SGLT1 and enhances intestinal mucosal glucose absorption capacity, which contributes to the understanding of intestinal physiologic function for patients with biliary duct obstruction and external biliary drainage.