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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.

Neuropilin-1 (NRP1) is a type I transmembrane glycoprotein that was originally found playing a role in neuronal axon guidance and embryonic angiogenesis. NRP1 can promote various aspects of tumorigenesis, such as angiogenesis, cell survival, migration, invasion and chemo-resistance, but little is known about the nanomechanica signature of NRP1 inregulating hepatocellular carcinoma(HCC) cells. Our study found that NRP1 was overexpressed in HCC and high expression of NRP1 was associated with poor prognosis in HCC patients.but there are no correlation between NRP1 and liver cirrhosis. In vivo and vitro experiments knockdown of NRP1 reduced proliferation and invasion of HCC. We quantified the Young's modulus, RMS residuals of HCC cell lines and normal liver cell line L02 using Atomic force microscopy (AFM). AFM data indicated that Young's modulus decreased in highly metastatic HCC cells, which is consistent with the verification results of NRP1 knockdown. Our analyses revealed that down regulation of NRP1 negatively correlated with increased Young's modulus in cells exhibiting differing degrees of invasiveness. Our molecular data showed that NRP1 activated AKT, ERK1/2, and epithelial-mesenchymal transition (EMT)-related signals in HCC cells and promoted tumor apoptosis and invasion. As an important biophysical property of cells, the membrane stiffness of HCC cells was further quantified by AFM force measurements, which indicated that these cells became much stiffer after NRP1 knockdown. These results collectively illuminated that Neuropilin-1 alterrated namomechanica signature in hepatocellular carcinoma cells and can be a potential therapeutic agent for HCC. Funding Statement: The present study was supported by the National Natural Science Fund of China (grant nos. 31570509 and 81702326), Science and Technology Major Project of Gansu province(1602FKDA001), the Talent innovation and Entrepreneurship Program of Lanzhou City (grant no. 2016-RC57) and the science and technology project of Chenguan District (2017SHFZ0014); Declaration of Interests: The authors declare no conflicts of interest. Ethics Approval Statement: The study was approved by the Zhongshan Hospital Ethics Committee and informed consent was obtained from each patient under Institutional Reviewer Board protocols. The study protocol conforms to the ethical guidelines of the 1975 Declaration of Helsinki as reflected in a prior approval by the institution's human research committee.

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.

This study investigates the biodegradation of crude oil by a mixed culture of bacteria isolated from the Qinghai-Tibet plateau using gas chromatography-mass spectrometer (GC-MS) and the gravimetric method. The results showed that a mixed culture has a stronger ability to degrade hydrocarbon than pure cultures. Once both Nocardia soli Y48 and Rhodococcus erythropolis YF28-1 (8) were present in a culture, the culture demonstrated the highest crude oil removal efficiency of almost 100% after 10 days of incubation at 20 °C. Moreover, further analysis of the degradation mechanisms used by the above strains, which revealed utilization of different n-alkane substrates, indicated the diversity of evolution and variations in different strains, as well as the importance of multiple metabolic mechanisms for alkane degradation. Therefore, it is concluded that a mixed culture of Y48 and YF28-1 (8) strains can provide a more effective method for bioremediation of hydrocarbon-contaminated soil in permafrost regions.

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

Our present study investigates the cellular and molecular inflammatory events in male albino rat arterial injury. Male albino rats were subjected to longitudinal incision and carotid artery clamping for the duration of 45 days. Heat shock protein (HSP) 27, HSP70, HSP47 and Nuclear Factor kappa B (NF-κB) expressions were determined by qPCR and Western blot method. The morphology of vessel wall alteration was studied by the light microscopy. The expression of NF-κB was found to be increased after ten days of carotid artery injury. The qPCR and Western blot analysis showed elevation in HSP47, HSP27, and HSP70 expression, ten days following the surgical injury. The neointima-formation and the media layer discontinuity were evidenced by light microscopy. The dendritic-like cells were in close contact with the lymphocytes. Our study reports that the surgical injury induces an inflammatory response through the increased NF-κB and HSPs expression.

The plant growth promoting rhizobacterium (PGPR) strain Bacillus amyloliquefaciens GB03, an important soil-borne bacterium, was shown to promote growth and abiotic stress tolerance in Arabidopsis thaliana as well as in some crop plants. This study aimed to evaluate the effects of GB03 on salt tolerance in Codonopsis pilosula, a traditional Chinese medicinal herb that is sensitive to salinity. Twenty-day-old seedlings of C. pilosula were either inoculated with GB03 or without it (as a control). At the same time, plants were treated with NaCl (0, 50, 100, or 150 mM) for 40 days. Growth parameters, photosynthetic indexes, malondialdehyde concentration, and leaf osmotic potential were measured after treatments. The result indicated that GB03 improved plant biomass of C. pilosula under salt conditions and improved the photosynthetic capacity by increasing net photosynthetic rate and stomatal conductance and decreasing intercellular CO2 concentration under both 0 and 50 mM NaCl. The bacterium strain also decreased leaf osmotic potential and peroxidation of membrane lipids that could help the plant adapt to saline environments. This study provides insights into the application of selected bacteria in the culture of important Chinese herbal plants under mild salinity.

Codonopsis pilosula (Franch.) Nannf. is a traditional Chinese herbal medicinal plant and a low-cost succedaneum for Panax ginseng and contains various bioactivity components. In this work, we first evaluated the effects of the inoculation of the plant growth-promoting rhizobacteria Bacillus amyloliquefaciens strain GB03 on growth and metabolite accumulation of C. pilosula. The results demonstrated that application of B. amyloliquefaciens GB03 significantly improved the growth of C. pilosula compared to DH5α, Luria broth medium, and water treatment, respectively. On the other hand, we observed that the content of lobetyolin, one of the most important secondary metabolites in C. pilosula, was obviously improved by inoculation of GB03 and almost reached twice that compared to the other three treatments. In addition, some amino acids of roots were elevated by GB03, although not significantly. In conclusion, B. amyloliquefaciens GB03 could induce positive effects on the growth and further stimulate accumulation of secondary metabolites in C. pilosula.

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

Cancer related fatigue significantly disrupts normal life for a portion of cancer survivors, and may persist for many years with cancer patients. Fatigue lasts longer than other treatment side-effects. There are increasing reports of using immunocyte therapy to treat cancer, and if it could be alternative relieve therapy in persons with cancer related fatigue is not so clear. Herein, we retrospectively evaluated the improvement of cancer related fatigue and safety of this therapy administered to cancer patients. 539 patients were treated with immunocyte therapy after conventional treatment. Methods: The delayed-type hyper-sensitivity (DTH) skin test was used to measure the immune response. Investigated into cancer related fatigue, physical strength, appetite and sleep as clinical efficacy. Results: The DTH skin tests showed that the positive cell-mediated cytotoxicity response rate was 82.7%. A large proportion of patients’ cancer related fatigue (80%); appetite (94.1%), sleeping status (96.6%) and metal state (92.9%) were improved. Adverse reactions were fever (35.1%), insomnia (21.4%), anorexia (15.7%), arthralgia (8%), rash (3.3%). DC (dendritic cell) therapy combined with CIK (cytokine-induced killer cell) was observed no toxicity in patients. Conclusion: This is the first time we reported combination of DC and CIK therapy to relieve the cancer related fatigue of patients after conventional therapy (surgery, radiotherapy, and chemotherapy). It may be an effective immunotherapy to reduce adverse reactions and a safety treatment. The data of CIK treatment time and CIK cell number provide strong support for further clinical treatment.

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.

Bacillus subtilis strain GB03 enhances growth and photosynthesis in the model plant Arabidopsis thaliana and several crop plants. In the present study, the effects of seed soaking with GB03 suspension culture and its volatile organic compounds on seed germination of Codonopsis pilosula (Franch.) Nannf. were investigated, and soil-grown C. pilosula seedlings were assayed to measure growth and photosynthetic capacity after soil inoculation with GB03. Both seed soaking with GB03 suspension culture and the presence of volatile organic compounds enhanced seed germination, especially seed germination vigour. GB03 significantly improved shoot and root length, branching, plant biomass (whole plant fresh and dry weight), leaf area and chlorophyll content in C. pilosula seedlings after 20, 40 and 60 days of soil inoculation. GB03 significantly enhanced transpiration rate, stomatal conductance and net photosynthetic rate, but decreased intercellular CO2 concentration. This study provides insight for the application of selected bacteria to improve biomass in Chinese herbal crops.