Quancheng Kan

miR-935 Promotes Liver Cancer Cell Proliferation and Migration by Targeting SOX7.

Hepatocellular carcinoma (HCC) is the most common cancer in the world. MicroRNAs (miRNAs) are a type of small noncoding RNA that can regulate the expression of target genes under physiological and pathophysiological conditions. Aberrant expression of microRNA-935 (miR-935) has been reported in cancer studies. However, its expression and mechanism in HCC remain unclear. In our study, we found that miR-935 was upregulated in liver cancer tissues and cells. Overexpression of miR-935 in liver cells promoted cell proliferation, tumorigenesis, and cell cycle progression, whereas inhibition of miR-935 reduced cell proliferation, tumorigenicity, and cell cycle progression. These changes in the properties of HCC cells were associated with upregulation of two well-known cellular G1/S transitional regulators: cyclin D1 and c-Myc. Additionally, we identified SOX7 as a direct target of miR-935. Overexpression of miR-935 inhibited SOX7 expression but promoted the levels of c-Myc and cyclin D1, which promotes cell proliferation and tumorigenesis; knockdown of miR-935 increased SOX7 level and inhibited c-Myc and cyclin D1 expression, whereas SOX7 silencing could promote cell proliferation, cell motility, and invasiveness in vitro. Our findings suggest that miR-935 represents a biomarker and a potential new target in HCC progression by suppressing SOX7 expression.

Galectin-1 Modulates the Survival and Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) Sensitivity in Human Hepatocellular Carcinoma Cells

Galectin-1 is a member of carbohydrate-binding proteins and plays critical roles in tumor growth and progression. It has been reported that galectin-1 is upregulated in human hepatocellular carcinoma (HCC) and facilitates HCC cell migration and invasion. In this study, the authors aimed to explore the effects of the knockdown of galectin-1 on HCC cell survival and sensitivity to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Human HCC cells were transfected with galectin-1-targeting small interfering RNA (siRNA) with or without 100 ng/mL TRAIL treatment and tested for apoptosis and gene expression changes. Cotransfection of Bcl-2- and survivin-expressing plasmids with galectin-1 siRNA was done, before TRAIL exposure, cell viability, and apoptosis were assessed. The authors found that siRNA-mediated downregulation of galectin-1 caused apoptosis in HCC cells, which was coupled with reduced Bcl-2 and survivin and increased Bax expression. Overexpression of Bcl-2 and survivin significantly blocked galectin-1 silencing-induced apoptosis of HCC cells. Knockdown of galectin-1 significantly enhanced TRAIL cytotoxicity against HCC cells, as determined by the MTT assay. Moreover, galectin-1 downregulation significantly induced apoptosis in TRAIL-treated HCC cells. Such effects were almost completely counteracted by the enforced expression of Bcl-2 and survivin. Taken together, these data first show that galectin-1 downregulation induces apoptosis in and augments TRAIL cytotoxicity to HCC cells largely through regulation of Bcl-2 and survivin expression. These findings provide a rationale for preclinical and clinical evaluation of targeting galectin-1 for improving TRAIL-based therapy against HCC.

Time‑dependent homeostasis between glucose uptake and consumption in astrocytes exposed to CoCl2 treatment

Hypoxia has been implicated in the pathology of the central nervous system during stroke. It also has a significant effect on the regulation of glucose transporters (GLUTs), and homeostasis between glucose uptake and consumption. CoCl2 is a hypoxia‑mimetic agent, and thus stabilizes the hypoxia‑inducible factor 1α (HIF‑1α) subunit and regulates GLUT genes. GLUT‑1 and GLUT‑3 are the most common isoforms of the GLUT family present in the brain, with the former primarily expressed in astrocytes and the latter in neurons under physiological conditions. However, it remains controversial whether GLUT‑3 is expressed in astrocytes. Additionally, it is unclear whether the regulation of GLUT‑1 and GLUT‑3, and glucose homeostasis, are affected by CoCl2 treatment in a time‑dependent manner. In the present study, mRNA and protein levels of GLUT‑1, GLUT‑3 and HIF‑1α in astrocytes were examined by reverse transcription‑quantitative polymerase chain reaction and western blot analysis, respectively. The intracellular glucose concentration, glycogen storage, ATP content, pyruvate concentration, lactate dehydrogenase (LDH) release activity and cell viability in astrocytes were also investigated. The observations of the current study confirmed that both protein and mRNA levels of GLUT‑1 and GLUT‑3 were elevated in a time‑dependent manner induced by CoCl2 treatment, followed by accumulation of HIF‑1α. Furthermore, in the early period of CoCl2 treatment (≤8 h at 100 µM), LDH release, ATP content, glycogen storage and cell viability remained unchanged, whereas intracellular pyruvate concentration increased and glucose concentration was reduced. However, in the later period of CoCl2 treatment (>8 h at 100 µM), LDH release and intracellular pyruvate concentration increased, while intracellular glucose concentration, ATP content and glycogen storage were reduced. This may be due to disruption of homeostasis and reduced cell viability. In conclusion, alteration in the expression levels of GLUT‑1 and GLUT‑3, and the homeostasis between glucose uptake and consumption were affected by CoCl2 treatment, in a time‑dependent manner, and may result in reduced energy production and cell viability in astrocytes.

The over-expression of FGFR4 could influence the features of gastric cancer cells and inhibit the efficacy of PD173074 and 5-fluorouracil towards gastric cancer

The aim was to investigate the function of fibroblast growth factor receptor 4 (FGFR4) in gastric cancer (GC) and explore the treatment value of agent targeted to FGFR4. Function assays in vitro and in vivo were performed to investigate the discrepancy of biological features among the GC cells with different expression of FGFR4. GC cells were treated with the single and combination of PD173074 (PD, an inhibitor of FGFR4) and 5-fluorouracil (5-Fu). The invasion ability were stronger, and the apoptosis rates were lower in MGC803 and BGC823 cells treated with FGFR4-LV5 (over-expression of FGFR4 protein) (Pu2009<u20090.05). The proliferation ability of GC cells is reduced when treated by the single and combination of 5-Fu and PD while that of the FGFR4-LV5 group was less inhibited compared with control group (Pu2009<u20090.05). The apoptosis rates are remarkably increased in GC cells treated with the single and combination of 5-Fu and PD (Pu2009<u20090.05). However, the apoptosis rate obviously is reduced in GC cells treated with FGFR4-LV5 compared with control group (Pu2009<u20090.05). The expression of PCNA and Bcl-XL is remarkably decreased, and the expression of Caspase-3 and cleaved Caspase-3 is obviously increased in GC cells treated with the single and combination of 5-Fu and PD. The tumor volumes of nude mice in FGFR4-LV5 group were much more increased (Pu2009<u20090.05). The over-expression of FGFR4 enhanced the proliferation ability of GC in vitro and in vivo. The combination of 5-Fu and PD exerted synergetic effect in weakening the proliferation ability and promoting apoptosis in GC cells, while the over-expression of FGFR4 might inhibit the efficacy of two drugs.

Time-dependent activity of Na+/H+ exchanger isoform 1 and homeostasis of intracellular pH in astrocytes exposed to CoCl2 treatment.

Hypoxia causes injury to the central nervous system during stroke and has significant effects on pH homeostasis. Na+/H+ exchanger isoformxa01 (NHE1) is important in the mechanisms of hypoxia and intracellular pH (pHi) homeostasis. As a well-established hypoxia-mimetic agent, CoCl2 stabilizes and increases the expression of hypoxia inducible factor‑1α (HIF-1α), which regulates several genes involved in pH balance, including NHE1. However, it is not fully understood whether NHE1 is activated in astrocytes under CoCl2 treatment. In the current study, pHi and NHE activity were analyzed using the pHi‑sensitive dye BCECF‑AM. Using cariporide (an NHE1‑specific inhibitor) and EIPA (an NHE nonspecific inhibitor), the current study demonstrated that it was NHE1, not the other NHE isoforms, that was important in regulating pHi homeostasis in astrocytes during CoCl2 treatment. Additionally, the present study observed that, during the early period of CoCl2 treatment (the first 2xa0h), NHE1 activity and pHi dropped immediately, and NHE1 mRNA expression was reduced compared with control levels, whereas expression levels of the NHE1 protein had not yet changed. In the later period of CoCl2 treatment, NHE1 activity and pHi significantly increased compared with the control levels, as did the mRNA and protein expression levels of NHE1. Furthermore, the cell viability and injury of astrocytes was not changed during the initial 8xa0h of CoCl2 treatment; their deterioration was associated with the higher levels of pHi and NHE1 activity. The current study concluded that NHE1 activity and pHi homeostasis are regulated by CoCl2 treatment in a time-dependent manner in astrocytes, and may be responsible for the changes in cell viability and injury observed under hypoxia-mimetic conditions induced by CoCl2 treatment.

-141C insertion/deletion polymorphism of the dopamine D2 receptor gene is associated with schizophrenia in Chinese Han population: Evidence from an ethnic group-specific meta-analysis.

Accumulate evidence has implicated dopamine D2 receptor gene polymorphisms in the etiology of schizophrenia. A single nucleotide polymorphism, −141C insertion/deletion (Ins/Del) (rs1799732), in the promoter region of the dopamine D2 receptor gene has been linked to schizophrenia; however, the data are inconclusive. This study investigated whether the −141C polymorphism is associated with the risk of schizophrenia in different ethnic groups by performing a meta‐analysis. A total of 24 case–control studies examining the association between −141C Ins/Del polymorphism and schizophrenia were identified according to established inclusion criteria. Significant association was revealed between −141C Ins/Del polymorphism and schizophrenia risk in dominant genetic model (Ins/Insu2009+u2009Ins/Del versus Del/Del) (odds ratiou2009=u20090.33, 95% confidence intervalu2009=u20090.14–0.81, zu2009=u20092.41, Pu2009=u20090.02) in Chinese Han but not in Caucasian, Japanese or India populations. Our results indicate that −141C Ins/Del polymorphism might be a susceptibility factor for schizophrenia in Chinese Han population.

Poly (ADP-ribose) polymerase- and cytochrome c-mediated apoptosis induces hepatocyte injury in a rat model of hyperammonia-induced hepatic failure

Hepatic failure (HF) is caused by several factors, which induce liver cell damage and dysfunction. However, the specific mechanism of HF remains to be fully elucidated. The present study aimed to investigate the underlying cause of hepatocyte injury and liver dysfunction. Liver cells were isolated from healthy female Sprague-Dawley rats, aged between 6 and 8 weeks, weighing ~230 g. The liver cells were cultured in RPMI-1640 medium containing 10% fetal bovine serum. An MTT assay was used to examine the inhibitory rate of liver growth in each group. Flow cytometric analysis was performed to detect liver cells undergoing apoptosis. The protein expression levels of poly (ADP-ribose) polymerase (PARP) and cytochrome c (Cyt C) were detected by western blotting. The level of calmodulin-dependent kinase (CaMK) was assessed using an ELISA. The results indicated that the growth inhibitory rate of rat liver cells was significantly increased following treatment with increasing concentrations of NH4Cl. The results of flow cytometric analysis demonstrated that the apoptotic rate in the BAPTA-acetoxymethyl ester group was significantly lower compared with the NH4Cl group (P<0.05). Treatment with NH4Cl increased the protein expression levels of PARP and Cyt C in the liver cells. The mRNA expression of CaMK decreased gradually following treatment with increasing concentrations of NH4Cl for 6, 12 and 24 h. The results suggested that hepatocyte injury and liver dysfunction may be caused by inducing apoptosis via the PARP and Cyt C pathways. Additionally, downregulation of CaMK may be associated with the apoptosis observed in hepatocyte injury.

Recombinant adenovirus expressing ICP47 gene suppresses the ability of dendritic cells by restricting specific T cell responses

Adenoviral vectors have been demonstrated to be one of the most effective vehicles to deliver foreign DNA into dendritic cells (DCs). However, the response of host immune systems against foreign gene products is a major obstacle to successful gene therapy. Infected cell protein 47 (ICP47) inhibits MHC Ⅰ antigen presentation pathway by binding to host transporter associated with antigen presentation (TAP), and thereby attenuates of specific cytotoxic T lymphocytes (CTLs) responses and evades the host immune clearance. This subject was designed to construct a recombinant adenovirus expressing His-tag-ICP47 fusion protein to investigate further the role of ICP47 in the elimination of transgene expression. Consequently, a recombinant adenovirus expressing the His-tag-ICP47 fusion protein was successfully constructed and it had the abilities of attenuating the stimulatory capacity of DCs by reducing the proliferation of lymphocytes and cytokine production of perforin compared with those of the r-track group and the control group. Our observations provide the first evidence of the regulation mechanism of ICP47 on DC-based immunotherapy for long-term persistence.

Gut microbiome analysis as a tool towards targeted non-invasive biomarkers for early hepatocellular carcinoma

Objective To characterise gut microbiome in patients with hepatocellular carcinoma (HCC) and evaluate the potential of microbiome as non-invasive biomarkers for HCC. Design We collected 486 faecal samples from East China, Central China and Northwest China prospectively and finally 419 samples completed Miseq sequencing. We characterised gut microbiome, identified microbial markers and constructed HCC classifier in 75 early HCC, 40 cirrhosis and 75 healthy controls. We validated the results in 56 controls, 30 early HCC and 45 advanced HCC. We further verified diagnosis potential in 18 HCC from Xinjiang and 80 HCC from Zhengzhou. Results Faecal microbial diversity was increased from cirrhosis to early HCC with cirrhosis. Phylum Actinobacteria was increased in early HCC versus cirrhosis. Correspondingly, 13 genera including Gemmiger and Parabacteroides were enriched in early HCC versus cirrhosis. Butyrate-producing genera were decreased, while genera producing-lipopolysaccharide were increased in early HCC versus controls. The optimal 30 microbial markers were identified through a fivefold cross-validation on a random forest model and achieved an area under the curve of 80.64% between 75 early HCC and 105 non-HCC samples. Notably, gut microbial markers validated strong diagnosis potential for early HCC and even advanced HCC. Importantly, microbial markers successfully achieved a cross-region validation of HCC from Northwest China and Central China. Conclusions This study is the first to characterise gut microbiome in patients with HCC and to report the successful diagnosis model establishment and cross-region validation of microbial markers for HCC. Gut microbiota-targeted biomarkers represent potential non-invasive tools for early diagnosis of HCC.

Role of sodium-hydrogen exchanger isoform 1 in regulating hepatocyte apoptosis induced by hyperammonaemia

BACKGROUNDnThe secondary injury theory of liver failure indicated that hyperammonaemia due to liver failure causes further deterioration of hepatocytes. Our previous studies have demonstrated that high blood ammonia levels may lead to hepatocyte apoptosis, as NH4Cl loading caused metabolic acidosis and an increase in sodium-hydrogen exchanger isoform 1 (NHE1). In this study, we established a hyperammonia hepatocyte model to determine the role of NHE1 in the regulation of hepatocyte apoptosis induced by NH4Cl.nnnMATERIALS AND METHODSnIn current studies, intracellular pH (pHi) and NHE1 activity were analyzed using the pHi-sensitive dye BCECF-AM. The results showed that intracellular pH dropped and NHE1 activity increased in hepatocytes under NH4Cl treatment. As expected, decreased pHi induced by NH4Cl was associated with increased apoptosis, low cell proliferation and ATP depletion, which was exacerbated by exposure to the NHE1 inhibitor cariporide. We also found that NH4Cl treatment stimulated PI3K and Akt phosphorylation and this effect was considerably reduced by NHE1 inhibition.nnnCONCLUSIONnThis study highlighted the significant role of NHE1 in the regulation of cell apoptosis induced by hyperammonaemia.