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BACKGROUND Hepatocellular carcinoma (HCC) is the third leading cause of cancer mortality worldwide. The gut microbiota can help maintain healthy metabolism and immunity. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a critical factor in promoting health and homeostasis; it promotes intestinal immunity, stimulates bone marrow precursors to generate macrophage colonies, and enhances the antibacterial and antitumor activity of circulating monocytes. As such, GM-CSF may protect against HCC development by regulating immunity as well as intestinal microecology. AIM To investigate the impact of GM-CSF on the gut microbiome and metabolic characteristics of HCC. METHODS Thirty-six male BALB/c nude mice were divided into three groups: Control (n = 10), HCC (n = 13), and HCC + GM-CSF (GM-CSF overexpression, n = 13). We utilized HCC cells to establish orthotopic transplantation tumor models of HCC with normal and over-expressing GM-CSF. Liver injury, immune inflammatory function and intestinal barrier function were evaluated. The fecal microbiome and metabolome were studied using 16S rRNA absolute quantification sequencing and gas chromatography-mass spectrometry. RESULTS GM-CSF overexpression significantly affected the gut microbiome of mice with HCC and resulted in a high abundance of organisms of the genera Roseburia, Blautia and Butyricimonass, along with a significant reduction in Prevotella, Parabacteroides, Anaerotruncus, Streptococcus, Clostridium, and Mucispirillum. Likewise, GM-CSF overexpression resulted in a substantial increase in fecal biotin and oleic acid levels, along with a prominent decrease in the fecal succinic acid, adenosine, fumaric acid, lipoic acid, and maleic acid levels. Correlation analysis revealed that the intestinal microbiota and fecal metabolites induced by GM-CSF were primarily involved in pathways related to reducing the inflammatory response, biotin metabolism, and intestinal barrier dysfunction. CONCLUSION GM-CSF can protect against HCC development by regulating immunity and modulating the abundance of specific intestinal microorganisms and their metabolites. This study provides new insights into the therapeutic approaches for HCC.

Background Neuropilin-1 (NRP-1) participates in cancer cell proliferation and metastasis as a multifunctional co-receptor by interacting with multiple signaling pathways. However, few studies have addressed the precise function and prognosis analysis of NRP1 in intrahepatic cholangiocarcinoma (ICC). We aimed to study the correlations between NRP1 and clinicopathological characteristics and NRP1 effect on ICC cell line functions. Methods NRP1 mRNA and its protein levels in human ICC tissues and cell lines were detected by IHC, qRT-PCR, and WB method. Transwell, wound healing, and CCK-8 assays were performed to verify the effects of NRP1 knockdown and overexpression on cell migration and proliferation capability. Results NRP1 proteins and mRNA levels increased in ICC tissues compared to those in paired adjacent non-tumor tissues. High NRP1 expression of ICC tissues was related to poor prognosis. NRP1 expression level was expected to be an independent prognosticator for overall survival and cumulative tumor recurrence, and was closely related to tumor number (P=0.047). Knockdown of NRP1 inhibited cell proliferation and migration capability of RBE cells in vitro, and NRP1 overexpression in 9810 cells accelerated proliferation and migration. Additionally, NRP1 may promote cell proliferation and migration in ICC via the FAK/PI3-K/AKT pathway. Conclusion As an oncogene, NRP1 may function as a candidate target and prognostic biomarker of value for ICC therapy.

A Gram-stain-positive, aerobic, non-motile and mycolic-acid-containing strain, designated Y48T, was isolated from soil contaminated by crude oil located in the northern margin of the Qaidam Basin. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain Y48T belongs to the genus Nocardia and is closely related to N. cummidelens DSM 44490T (99.0 % similarity), N. soli DSM 44488T (99.0 %), N. lasii 3C-HV12T (98.9 %), N. salmonicida NBRC 13393T (98.6 %), N. ignorata NBRC 108230T (98.6 %) and N. coubleae NBRC 108252T (98.6 %). The average nucleotide identity and DNA–DNA hybridization values between strain Y48T and the reference strains were 75.9–84.5 and 27.5–29.0 %, respectively, values that were below the thresholds for species delineation. Chemotaxonomic analysis indicated that the major fatty acids of strain Y48T were C16 : 0, summed feature 3 (C16 : 1 ω6c/C16 : 1 ω7c), C18 : 1 ω9c and C18 : 0 10-methyl (TBSA). The respiratory quinone was MK-8(H4, ω-cycl). The polar lipid profile was composed of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannoside, two glycolipids and three unidentified lipids. The cell-wall hydrolysates contained meso-diaminopimelic acid, with ribose, arabinose, glucose and galactose as whole-cell sugars. A combination of 16S rRNA gene sequence analysis, and phenotypic and chemotaxonomic characterizations demonstrated that strain Y48T represents a novel species of the genus Nocardia , for which the name Nocardia mangyaensis sp. nov. is proposed. The type strain is Y48T (=JCM 32795T=CGMCC 4.7494T).

Background Intestinal microbial dysbiosis is involved in liver disease pathogenesis. However, its role in primary liver cancer (PLC), particularly in hepatocarcinogenesis remains unclear. The present study aimed to study the changes in intestinal flora at various stages of PLC and clarify the relationship between intestinal microbes and PLC. Methods Twenty-four patients with PLC (PLC group), 24 patients with liver cirrhosis (LC group), and 23 healthy control individuals (HC group) were enrolled from October 2016 to October 2017. Stool specimens of the participants were collected and the genomic DNA of fecal bacteria was isolated. High-throughput pyrosequencing of 16S rDNA was used to identify differences in gut bacterial diversity among HC, LC, and PLC groups. We also analyzed the relationship between clinical factors and intestinal microorganisms in LC and PLC groups. Results Diversity of Firmicutes tended to decrease from the HC to LC and PLC groups at the phylum level. Among species, Enterobacter ludwigii displayed an increasing trend in the PLC group, wherein the relative abundance of Enterobacter ludwigii in the PLC group was 100 times greater than that in the HC and LC groups. The ratio of Firmicutes/Bacteroidetes was significantly decreased with the disease progression. In addition, the linear discriminant analysis effect size method indicated that Clostridia were predominant in the gut microbiota of the HC group, whereas Enterococcaceae, Lactobacillales, Bacilli and Gammaproteobacteria may be used as diagnostic markers of PLC. Redundancy analysis showed a correlation between intestinal microbial diversity and clinical factors AST, ALT, and AFP. Veillonella showed a significant positive correlation with AFP in the PLC group, whereas Subdoligranulum showed a negative correlation with AFP. Conclusions This study indicates that dysbiosis of the gut microbiota might be involved in PLC development and progression.

A possible association between multiple drug resistance 1 gene (MDR1) polymorphisms and the risk of developing hepatocellular carcinoma (HCC) is currently under debate, and evidence from various epidemiological studies has yielded controversial results. To derive a more precise estimation of the association between MDR1 polymorphisms and HCC risk, the present meta-analysis was performed. A total of 8 studies containing 11 cohorts with 4407 cases and 4436 controls were included by systematic literature search of EMBASE, PubMed, Web of Science, and CNKI. All polymorphisms were classified as mutant/wild-type alleles. In particular, the variation type, functional impact, and protein domain location of the polymorphisms were assessed and used as stratified indicators. The pooled odds ratio (OR) with 95 % confidence interval (CI) was calculated to evaluate the association. Overall, our results suggested that the mutant alleles of the MDR1 gene were associated with a significantly increased risk for HCC under all genetic models (allelic model: OR = 1.28, 95 % CI = 1.20–1.36, P

An increasing body of evidence suggests that down regulation or deletion of HECT domain and ankyrin repeat containing E3 ubiquitin protein ligase 1 (HACE1) gene plays an important role in the occurrence, invasion and metastasis process in many human malignancies and is closely related to prognosis. However, sparse evidence exists concerning the precise function and clinical significance of HACE1 in hepatocellular carcinoma (HCC). In the present study, we investigated the expression pattern of HACE1 in HCC tissues and cell lines, and determined the potential functions of HACE1 in HCC cell lines and evaluated the relationships between HACE1 expression and clinicopathological characteristics. Protein and mRNA expression levels of HACE1 in human HCC tissues and cell lines were examined by western blot analysis, quantitative real-time polymerase chain reaction and immunohistochemical (IHC) analyses. IHC was used to analyze the correlations between HACE1 expression and clinicopathological features. HACE1 was upregulated in SMCC7721 cells by transfection with pcDNA3.1-HACE1 and Huh7 cells were transfected with siRNA targeting HACE1 for downregulation. Cell Counting Kit-8, Transwell and wound healing assays were performed to investigate the effects of the overexpression and knockdown of HACE1 on cellular proliferation and migration. The results revealed that HACE1 expression was lower in the HCC tissues and cell lines at the mRNA and protein levels compared to levels noted in the matched non-tumor tissues and the normal liver cell line L02. Knockdown of HACE1 in Huh7 cells accelerated cell proliferation and migration (P

Hepatocellular carcinoma (HCC) is the third-most lethal cancer worldwide. Understanding the molecular pathogenesis of HCC recurrence and metastasis is the key to improve patients’ prognosis. In this study, we report that protein tyrosine phosphatase receptor S (PTPRS) is significantly down-regulated in nearly 80% of HCCs, and its expression negatively correlates with aggressive pathological features, such as larger tumor size and advanced stage. In addition, PTPRS deficiency is independently associated with shorter survival and increased recurrence in patients, although 16.7% of HCCs show intratumor heterogeneous expression of PTPRS. Restoration of wild-type, but not mutant, PTPRS expression significantly inhibits HCC cell migration and invasion in vitro as well as lung metastasis in vivo, whereas knockdown of its expression significantly promotes invasion and metastasis. Notably, PTPRS-regulated HCC invasiveness is accompanied by typical changes of epithelial-mesenchymal transition (EMT). Moreover, PTPRS forms a complex with epithermal growth factor receptor (EGFR) and regulates its tyrosine residues’ phosphorylation. Ectopic expression of EGFR reverses the metastasis-inhibiting effects of PTPRS, whereas silencing of EGFR or inhibiting phosphorylation of key molecules in EGFR downstream pathways reinhibits EMT and metastasis caused by PTPRS down-regulation. Meanwhile, promoter hypermethylation of PTPRS is frequently detected in HCC samples and cell lines. Treatment with a demethylation agent, 5-aza-2′-deoxycytidine, recovers PTPRS expression in a dose-dependent manner. Conclusions: Epigenetic inactivation of PTPRS may increase phosphorylation and activity of EGFR signaling to promote EMT and metastasis in HCC. (Hepatology 2015;62:1201-1214)

Soil salinity is an increasingly serious problem worldwide that reduces agricultural output potential. Selected beneficial soil bacteria can promote plant growth and augment tolerance to biotic and abiotic stresses. Bacillus subtilis strain GB03 has been shown to confer growth promotion and abiotic stress tolerance in the model plant Arabidopsis thaliana. Here we examined the effect of this beneficial soil bacterium on salt tolerance in the legume forage crop, white clover. Plants of white clover (Trifolium repens L. cultivar Huia) were grown from seeds with or without soil inoculation of the beneficial soil bacterium Bacillus subtilis GB03 supplemented with 0, 50, 100, or 150 mM NaCl water into soil. Growth parameters, chlorophyll content, malondialdehyde (MDA) content and osmotic potential were monitored during the growth cycle. Endogenous Na(+) and K(+) contents were determined at the time of harvest. White clover plants grown in GB03-inoculated soil were significantly larger than non-inoculated controls with respect to shoot height, root length, plant biomass, leaf area and chlorophyll content; leaf MDA content under saline condition and leaf osmotic potential under severe salinity condition (150 mM NaCl) were significantly decreased. Furthermore, GB03 significantly decreased shoot and root Na(+) accumulation and thereby improved K(+)/Na(+) ratio when GB03-inoculated plants were grown under elevated salt conditions. The results indicate that soil inoculation with GB03 promotes white clover growth under both non-saline and saline conditions by directly or indirectly regulating plant chlorophyll content, leaf osmotic potential, cell membrane integrity and ion accumulation.