Brachyury promotes proliferation and migration of hepatocellular carcinoma via facilitating the transcription of NCAPG2.

Hepatocellular carcinoma (HCC) has a poor prognosis because of its limited drug responses in clinical trials. Therefore, it is crucial to clarify the molecular mechanisms of HCC progression to identify new diagnostic markers and therapeutic targets. Here, we report that brachyury, which regulates the gene encoding the non-SMC condensin II complex subunit G2 (NCAPG2), promotes tumorigenesis in HCC. Knockdown of brachyury led to inhibition of cancer progression in vitro and in vivo. Chromatin immunoprecipitation-sequencing data indicated that the oncogene NCAPG2 is a direct target of brachyury. Furthermore, NCAPG2 knockdown inhibited the proliferation and migration of HCC cells and attenuated brachyury-induced tumorigenesis. Overexpression and decreased DNA methylation of NCAPG2 were associated with a poor prognosis, and NCAPG2 was positively correlated with various immune cell infiltrates, cancer-associated fibroblasts, and immune checkpoint molecule expression levels in the tumor microenvironment. Moreover, the effectiveness of immune checkpoint blockade was decreased in the high NCAPG2 expression group. Together, these findings demonstrated a coregulatory effect of the brachyury/NCAPG2 axis during HCC progression.

Modafinil ameliorated pancreatic injury and inflammation through upregulating SNIP1.

Acute pancreatitis (AP) is the inflammatory response of the exocrine pancreas to various causes. Modafinil has significant anti-inflammation and anti-oxidation effects. No experiment has assessed the effects of modafinil on AP. Thus, the study aims to study the effects of modafinil on AP and its potential mechanism in vivo and vitro. 5% sodium taurocholate was retrograde injected into pancreatic duct to establish AP rat model. The severity of AP was detected by HE staining, serum amylase and lipase levels. The inflammation, oxidative stress and apoptosis were detected separately by ELISA, MDA and SOD kits, tunnel staining and Western blotting in rats. Besides, SNIP1 expression was analyzed by qPCR and Western blotting. In vivo, AR42J cells were stimulated by cerulein and lipopolysaccharide to establish AP cell model. Flow cytometry examined cell apoptosis. After the plasmids silencing SNIP1 were transfected into AP cells, the inhibitory effects of modafinil on inflammation, oxidative stress and apoptosis were significantly reversed. The results indicated that modafinil showed significant curative and therapeutic effects by regulating SNIP1 level.

Sinensetin flavone exhibits potent anticancer activity against drug-resistant human gallbladder adenocarcinoma cells by targeting PTEN/PI3K/AKT signalling pathway, induces cellular apoptosis and inhibits cell migration and invasion.

PURPOSE: The main focus of the current research work was to unveil the anticancer activity of the naturally occurring Sinensetin flavone against aggressive gall bladder cancer adenocarcinoma (GBAC) TJ-GBC2 cell line. Its effect of inducing apoptosis mediated via targeting PTEN/PI3K/AKT signalling pathway were also examined along with cell migration and invasion. METHODS: Cell proliferation was tested by MTT cell viability assay. Fluorescence microscopy was utilized to carry out apoptosis related studies via DAPI staining along with flow cytometry using annexin V/propidium iodide (PI) assay. Further, western blotting analysis was carried out to examine the effects of Sinensetin on the expressions of apoptosis-related proteins and Bax Bcl-2 along with PTEN/PI3K/AKT signalling pathway. The impact of the test molecule on cell migration and invasion was studied through wound healing assay and transwell cell invasion assay respectively. RESULTS: The results showed that Sinensetin treatment caused a significant retardation in cell viability, in a dose-dependent fashion. DAPI staining assay and annexin V/PI assay revealed that the cell viability of GBC cells was retarded due to induction of apoptosis. It was also associated with downregulation of Bcl-2 and upregulation of Bax levels. Further, wound healing assay and transwell cell invasion assay revealed that cell migration as well as cell invasion of cancer gallbladder cells was decreased in a concentration-dependent fashion. It was further seen that Sinensetin treatment resulted in inhibition of matrix metalloproteinase (MMP)-2 and enhancement of MMP-9 protein expressions. Results also showed that the tested molecule had the potential to inhibit PTEN/PI3K/AKT signalling pathway. CONCLUSION: In conclusion, the current study indicated that Sinensetin flavone has the potential to be developed as a candidate drug against gallbladder adenocarcinoma provided more toxicological and in vivo studies are carried out.

Axin1 inhibits proliferation, invasion, migration and EMT of hepatocellular carcinoma by targeting miR-650.

Hepatocellular carcinoma (HCC) is one of the most common malignant tumors with a high mortality rate and low survival rate. This study was designed to explore a novel molecular with high sensitivity and specificity, which can be applied in early diagnosis and therapeutic evaluation of HCC. The current study aims to investigate the effect and important role of Axin1 on cell proliferation, invasion, migration and epithelial-mesenchymal transition (EMT) in hepatocellular carcinoma. qRT-PCR results showed lower Axin1 expression level and higher miR-650 expression level in HCC. Luciferase reporter assay was carried out to verify the negative correlation between Axin1 and miR-650 mRNA levels. CCK-8 assay results showed that the cell proliferation ability was significantly suppressed by Axin1 overexpression in SK-HEP-1 cells. The results in wound healing assay uncovered that cell migration ability was markedly suppressed by Axin1 overexpression. The results in trans-well invasion assay showed that Axin1 overexpression caused decreased invasive ability in SK-HEP-1 cells. The WB results showed that the protein level of E-cad was significantly increased and the protein levels of N-cad, vimentin and snail were obviously reduced following Axin1 overexpression. Whereas, the suppressive effects on cell proliferation, migration, invasion and EMT caused by Axin1 overexpression were abolished by miR-650 mimic. All the results in the current study confirmed the truth that Axin1 overexpression could suppress cell proliferation, migration, invasion and EMT by downregulating miR-650 expression.

Role of miR-18a and miR-25 disruption and its mechanistic pattern in progression of liver cancer.

This study examined the molecular mechanisms underlying the roles of the microRNAs miR-18a and miR-25 in the progression of human liver cancer. Liver cancer biopsies obtained from early-stage liver cancer patients were examined by qRT-PCR and Northern blotting to examine the expression of miR-18a and miR-25. Both microRNAs were overexpressed in mouse primary hepatocytes following transfection of the cells with vectors encoding the microRNAs. An analysis of biopsy samples from liver cancer patients indicated that both miR-18a and miR-25 were overexpressed during the early stages of liver cancer. Further, qRT-PCR and Northern blotting confirmed that both of these microRNAs play crucial roles in the progression of liver cancer. Our findings clearly indicate that miR-18a and miR-25 can be used as prognostic biomarkers for early-stage liver cancer. Hence, miR-18a and miR-25 may have value as prognostic indicators and may facilitate the development of novel therapeutics for liver cancer.

miR-1470 regulates cell proliferation and apoptosis by targeting ALX4 in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common malignant tumors worldwide. MicroRNAs (miRNAs) have been proven to play essential roles in different cancers, including HCC. The current study was mainly focused on the role of miR-1470 in HCC progression. METHODS: Quantitative real-time PCR (qRT-PCR) was performed to detect the expression levels of miR-1470 and Aristaless-like homeobox-4 (ALX4). The CCK-8 and EdU assays were used to examine cell proliferation. Flow cytometric analysis was used to elucidate the cell cycle and cell apoptosis. A xenograft tumor assay was carried out to verify the effect of miR-1470 on tumor formation in vivo. RESULTS: According to the qRT-PCR assay, miR-1470 was proven to be overexpressed in HCC. As shown by the CCK-8 assay, EdU assay and flow cytometric analysis, miR-1470 overexpression promoted cell proliferation and inhibited cell apoptosis. ALX4 was proven via a dual luciferase reporter assay to be a downstream target gene of miR-1470. ALX4 was downregulated in HCC. The results of a rescue assay revealed that miR-1470 had an oncogenic role in HCC by regulating ALX4. CONCLUSION: miR-1470 exhibits an oncogenic role in HCC by targeting ALX4. The data from our study may provide novel insight for the identification of new biomarkers and treatment strategies for HCC.

Comparison on the Efficacy of Three Duct Closure Methods after Laparoscopic Common Bile Duct Exploration for Choledocholithiasis.

BACKGROUND Laparoscopic common bile duct exploration (LCBDE) is currently the best approach for complex cases of choledocholithiasis or the cases of endoscopic retrograde cholangiopancreatography (ERCP) failure. Nevertheless, there is no clear consensus on the optimal duct closure method after LCBDE. The purpose of this study was to evaluate the efficacy of 3 duct closure methods after LCBDE for choledocholithiasis. MATERIAL AND METHODS In this analysis, 189 patients with choledocholithiasis underwent LCBDE between June 2014 and December 2018. According to different duct closure methods, these patients were divided into T-tube drainage (TTD) group (n=66), common suture group (n=64) and barbed suture group (n=59). The operation time, suturing time, amount of intraoperative bleeding, tube-carried time, length of stay (LOS), hospitalization costs, pre- and post-operative common bile duct (CBD) diameters were all compared among the 3 groups. Six months after discharge, the incidence of complications and recurrent stones was observed. RESULTS The operation time, suturing time, and amount of intraoperative bleeding in barbed suture group were both significantly less than those in the common suture group and the TTD group (P<0.01). When compared with the TTD group, the suturing time, tube-carried time, and LOS were decreased markedly in the common suture group and the barbed suture group (P<0.01). The post-operative CBD diameters in the 3 groups were all significantly larger than the pre-operative CBD diameters (P<0.01). There was no statistical significance among the 3 groups regarding the incidence of complications and recurrent stones (P>0.05). CONCLUSIONS Barbed suture shortened the suturing time, operation time, tube-carried time, and LOS, and lessened the amount of intraoperative bleeding in patients with choledocholithiasis after LCBDE. It was more effective than the common suture and TTD.

Overexpression of RAB34 correlates with poor prognosis and tumor progression in hepatocellular carcinoma.

RAB34, a protein belonging to the RAB family, is involved in protein transport, repositioning of lysosomes and activation of micropinocytosis. However, few studies have reported its function in human epithelial cancers. Immunohistochemistry (IHC) and western blotting were used to detect expression of RAB34 at the tissue and cell levels. Cell Counting Kit-8 (CCK-8), EDU assay and flow cytometry were used for analyzing cell proliferation. Transwell and scratch wound healing assays were used for assessing cell migration ability. Western blotting was used for detecting expression of E-cadherin and N-cadherin. In the present study, we found that both DNA copy and protein level of RAB34 were upregulating in human hepatocellular carcinoma (HCC) tissues when compared with that in adjacent tissues. Analysis of the correlation between RAB34 expression and clinicopathological features showed that patients with overexpression of RAB34 consistently had large tumor size, vessel invasion and poor tumor grade. Furthermore, overall survival analysis showed that patients with upregulated expression of RAB34 were associated with poor prognosis. Moreover, cell function experiments showed that suppression of RAB34 led to a lower proliferation rate and migration ability. In addition, this phenomenon may be attributed to cell cycle phase G1 arrest and mesenchymal-epithelial transition under condition of RAB34 suppression. The present study demonstrated that RAB34 plays an important role in the initiation and progression of HCC. Our results suggest a new therapeutic target for the clinical treatment of HCC.

Inhibition of STAT3/cyclinD1 pathway promotes chemotherapeutic sensitivity of colorectal caner.

BACKGROUND: Chemotherapeutic resistance indicated the poor prognosis of colorectal cancer. OBJECTIVE: Our study aimed to investigate the role of STAT3/cyclinD1 pathway in the chemotherapeutic resistance of colorectal cancer. METHODS: We firstly measured the expression of cyclinD1 in the colorectal cancer tissues using immunohistochemistry in tissue microarray. Then cell viability and apoptosis were investigated in the HT-29 cell lines dealing with recombinant lentivirus and shRNA to increase or decrease cyclinD1 expression. Furthermore, luciferase and ChIP assays were applied to investigate whether STAT3 regulated cyclinD1 expression by binding to its promoter. Finally, we determined whether inhibition of STAT3 could decrease cyclinD1 and increase the chemotherapy sensitivity. RESULTS: CyclinD1 expression was significantly increased in the cancer cells and high level of cyclinD1 indicated the poor prognosis. Inhibition of cyclinD1 decreased the cell viability assessed by MTT and increased rate of apoptosis when exposed to 5-FU treatment while overexpression of cyclinD1 showed the reverse effect. ChIP assay showed that STAT3 directly bind to cyclinD1 promoter. Subclone of full promoter of cyclinD1 into pGL4 increased the luciferase activity while delete or mutation of any of STAT3 binding sites resulted in reductions of luciferase activity. Inhibition of STAT3 decreased cyclinD1 expression to decrease the cell viability and increase rate of apoptosis when exposed to 5-FU treatment. CONCLUSIONS: Inhibition of STAT3/cyclinD1 pathway increased the sensitivity of colorectal cancer cell to chemotherapy.