Chemical characteristics of dissolved organic matter in effluent from sludge alkaline fermentation liquid-fed sequencing batch reactors.

Sludge alkaline fermentation liquid (SAFL) is a promising alternative to acetate for improving biological nitrogen removal (BNR) from wastewater. SAFL inevitably contains some refractory compounds, while the characteristics of dissolved organic matter (DOM) in effluent from SAFL-fed BNR process remain unclear. In this study, the molecular weight distribution, fluorescent composition and molecular profiles of DOM in effluent from SAFL and acetate-fed sequencing batch reactors (S-SBRs and A-SBRs, respectively) at different hydraulic retention time (12 h and 24 h) was comparatively investigated. Two carbon sources resulted in similar effluent TN, but a larger amount of DOM, which was bio-refractory or microorganisms-derived, was found in effluent of S-SBRs. Compared to acetate, SAFL increased the proportion of large molecular weight organics and humic-like substances in effluent DOM by 74.87%-101.3% and 37.52%-48.35%, respectively, suggesting their bio-refractory nature. Molecular profiles analysis revealed that effluent DOM of S-SBRs exhibited a more diverse composition and a higher proportion of lignin-like molecules. Microorganisms-derived molecules were found to be the dominant fraction (71.51%-72.70%) in effluent DOM (<800 Da) of S-SBRs. Additionally, a prolonged hydraulic retention time enriched Bacteroidota, Haliangium and unclassified_f_Comamonadaceae, which benefited the degradation of DOM in S-SBRs. The results help to develop strategies on reducing effluent DOM in SAFL-fed BNR process.

Fabrication of a superhydrophobic surface by straightforward immersion for copperplate artwork protection.

With the background of contemporary art, using comprehensive materials to create artworks is becoming more and more common. The new era of digital image-based copperplate artworks, using photosensitive lithography, has given traditional art forms new life and greater popularity in the digital age. However, the patterns and textures of the works created by the new techniques are generally shallow, and the copper surface is easily damaged and loses its aesthetic value, which makes it a practical problem to protect such works more effectively. In this paper, a facile method is adopted, wherein a superhydrophobic film is constructed on the surface of copperplate images by straightforward immersion in (heptadecafluoro-1,1,2,2-tetradecyl)trimethoxysilane (FAS-17) solution to achieve the anticorrosive protection of copperplate artworks. The hydrophobicity of the copper surface was analyzed using an instrument that measures contact angles. The superhydrophobic surface morphology and composition were analyzed with a scanning electron microscope coupled with an energy-dispersive spectrometer, and the corrosion resistance was analyzed using an electrochemical workstation. A systematic study is presented on the effect of the immersion time in FAS-17 and the concentration of FAS-17, and the optimal preparation conditions of the superhydrophobic film were determined, which means that the copper substrates were immersed in 0.7 mol L-1 FAS-17 for 40 min. After the treatment of the surface to make it superhydrophobic, the contact angle and the corrosion inhibition efficiency of the copperplate etching surface reached 161.2° and 95.7%, respectively. The results show that the superhydrophobic film was successfully prepared on the surface of the artwork based on copper, which can effectively improve the corrosion resistance and is beneficial for the long-term protection of artwork.

Magnetic resonance imaging DTI-FT study on schizophrenic patients with typical negative first symptoms.

Magnetic resonance imaging (MRI) with diffusion-tensor imaging (DTI) together with a white matter fiber tracking (FT) technique was used to assess different brain white matter structures and functionalities in schizophrenic patients with typical first negative symptoms. In total, 30 schizophrenic patients with typical first negative symptoms, comprising an observation group were paired 1:1 according to gender, age, right-handedness, and education, with 30 healthy individuals in a control group. Individuals in each group underwent routine MRI and DTI examination of the brain, and diffusion-tensor tractography (DTT) data were obtained through whole brain analysis based on voxel and tractography. The results were expressed by fractional anisotropy (FA) values. The schizophrenic patients were evaluated using a positive and negative symptom scale (PANSS) as well as a Global Assessment Scale (GAS). The results of the study showed that routine MRIs identified no differences between the two groups. However, compared with the control group, the FA values obtained by DTT from the deep left prefrontal cortex, the right deep temporal lobe, the white matter of the inferior frontal gyrus and part of the corpus callosum were significantly lower in the observation group (P<0.05). The PANSS positive scale value in the observation group averaged 7.7±1.5, and the negative scale averaged 46.6±5.9, while the general psychopathology scale averaged 65.4±10.3, and GAS averaged 53.8±19.2. The Pearson statistical analysis, the left deep prefrontal cortex, the right deep temporal lobe, the white matter of the inferior frontal gyrus and the FA value of part of the corpus callosum in the observation group was negatively correlated with the negative scale (P<0.05), and positively correlated with GAS (P<0.05). In conclusion, a decrease in the FA values of the left deep prefrontal cortex, the right deep temporal lobe, the white matter of the inferior frontal gyrus and part of the corpus callosum may be associated with schizophrenia with typical first negative symptoms and the application of MRI DTI-FT can improve diagnostic accuracy.

Transdermal solid delivery of epigallocatechin-3-gallate using self-double-emulsifying drug delivery system as vehicle: Formulation, evaluation and vesicle-skin interaction.

The present study investigated a self-double-emulsifying drug delivery system loaded with epigallocatechin-3-gallate to improve epigallocatechin-3-gallate skin retention. The long chain solid lipids (cetostearyl alcohol) and macadamia oil were utilized as a carrier to deliver the bioactive ingredient. Response surface methodology was used to optimize the formulation, and the solid lipid to total lipid weight ratio, concentration of epigallocatechin-3-gallate and hydrophilic surfactant on skin retention were found to be the principal factors. The optimum formulation with high encapsulation efficiency (95.75%), self-double-emulsification performance (99.58%) and skin retention (87.24%) were derived from the fitted models and experimentally examined, demonstrating a reasonable agreement between experimental and predicted values. Epigallocatechin-3-gallate-self-double-emulsifying drug delivery system was found to be stable for 3 months. Transdermal studies could explain a higher skin diffusion of epigallocatechin-3-gallate from the self-double-emulsifying drug delivery system compared with EGCG aqueous solution. In vitro cytotoxicity showed that epigallocatechin-3-gallate-self-double-emulsifying drug delivery system did not exert hazardous effect on L929 cells up to 1:10.

Targeting SHCBP1 inhibits cell proliferation in human hepatocellular carcinoma cells.

Src homology 2 domain containing (SHC) is a proto-oncogene which mediates cell proliferation and carcinogenesis in human carcinomas. Here, the SHC SH2-domain binding protein 1 (SHCBP1) was first established to be up-regulated in human hepatocellular carcinoma (HCC) tissues by array-base comparative genome hybridization (aCGH). Meanwhile, we examine and verify it by quantitative real-time PCR and western blot. Our current data show that SHCBP1 was up-regulated in HCC tissues. Overexpression of SHCBP1 could significantly promote HCC cell proliferation, survival and colony formation in HCC cell lines. Furthermore, knockdown of SHCBP1 induced cell cycle delay and suppressed cell proliferation. Furthermore, SHCBP1 could regulate the expression of activate extracellular signal-regulated kinase 1/2 (ERK1/2) and cyclin D1. Together, our findings indicate that SHCBP1 may contribute to human hepatocellular carcinoma by promoting cell proliferation and may serve as a molecular target of cancer therapy.

Autophagy inhibition suppresses pulmonary metastasis of HCC in mice via impairing anoikis resistance and colonization of HCC cells.

Metastasis is one of the main causes of poor prognosis for hepatocellular carcinoma (HCC), which has been linked to cell-death resistance. Autophagy is an important survival mechanism under conditions of cell stress. We hypothesized that autophagy may play a role in HCC metastasis due to its prosurvival effect. Highly metastatic HCC cell lines with stable autophagy inhibition were established via lentivirus-mediated silencing of BECN1 and ATG5 genes. Mouse models of pulmonary metastasis were then developed using the cells with or without autophagy inhibition. The analysis of lung metastasis by histopathological examination and small animal imaging showed that autophagy inhibition significantly decreased the incidence of pulmonary metastases in vivo. Further invasion, migration, detachment, lung colonization, and epithelial-mesenchymal transition (EMT) assays indicated that autophagy inhibition did not affect cell invasiveness, migration or EMT but attenuated the anoikis-resistance and lung colonization of HCC cells. Investigation of the molecular mechanisms underlying showed that the autophagy-inhibition-mediated anoikis-resistance attenuation was associated with the regulation of apoptotic signaling. As autophagy inhibition was shown to be able to suppress HCC metastasis, an autophagy-based HCC tissue-specific target therapy system (AFP-Cre/LoxP-shRNA) was constructed. In vitro and in vivo analyses showed that the system was able to efficiently inhibit autophagy of HCC cells and tissue in a tissue-specific manner. Further in vivo metastasis assay showed that intratumoral administration of the system could significantly suppress lung metastasis. Together, our findings suggest that autophagy may be involved in HCC metastasis through facilitating anoikis resistance and lung colonization of HCC cells. Autophagy-based HCC tissue-specific target therapy may be a new strategy for the management of HCC metastasis.

α-Fetoprotein promoter-driven Cre/LoxP-switched RNA interference for hepatocellular carcinoma tissue-specific target therapy.

BACKGROUND: RNA interference (RNAi) has recently emerged as a potential treatment modality for hepatocellular carcinoma (HCC) therapy, but the lack of cellular targets and sustained efficacy limits its application. The purpose of this study is to develop an HCC tissue-specific RNAi system and investigate its possibility for HCC treatment. METHODS: Two different HCC-specific RNAi systems in which therapeutic miRNA or shRNA against target gene (Beclin 1) was directly or indirectly driven by alpha-fetoprotein promoter (AFP-miRNA and AFP-Cre/LoxP-shRNA) were constructed. Human HCC cell lines (HepG2, Hep3B and HCCLM3) and non-HCC cell lines (L-02, Hela and SW1116) were infected with the systems. The effectiveness and tissue-specificity of the systems were examined by Q-PCR and western blot analysis. The efficacy of the systems was further tested in mouse model of HCC by intravenous or intratumoral administration. The feasibility of the system for HCC treatment was evaluated by applying the system as adjuvant therapy to enhance sorafenib treatment. An AFP-Cre/LoxP-shRNA system targeting Atg5 gene (AFP-Cre/LoxP-shRNA-Atg5) was constructed and its efficacy in sensitizing HCC cells (MHCC97L/PLC) to sorafenib treatment was examined by apoptosis assay in vitro and tumorigenesis assay in vivo. RESULTS: The AFP-miRNA system could silence target gene (Beclin 1) but required a high titer which was lethal to target cells. The AFP-Cre/LoxP-shRNA system could efficiently knockdown target gene while maintain high HCC specificity. Intratumoral injection of the AFP-Cre/LoxP-shRNA system could efficiently silence target gene (Beclin 1) in vivo while intravenous administration could not. The AFP-Cre/LoxP-shRNA system target Atg5 gene could significantly sensitize MHCC97L/PLC cells to sorafenib-induced apoptosis in vitro and tumor growth suppression in vivo. CONCLUSIONS: An efficient HCC tissue-specific RNAi system (AFP-Cre/LoxP-shRNA) was successfully established. The system provides a usable tool for HCC-specific RNAi therapy, which may serve as a new treatment modality for HCC.

Up-regulation of NICE-3 as a novel EDC gene could contribute to human hepatocellular carcinoma.

The epidermal differentiation complex (EDC) contains a large number of gene products which are crucial for the maturation of the human epidermis and can contribute to skin diseases, even carcinogenesis. It is generally acepted that activation of oncogenes and/or inactivation of tumor suppressor genes play pivotal roles in the process of carcinogenesis. Here, NICE-3, a novel EDC gene, was found to be up-regulated in human hepatocellular carcinoma (HCC) by quantitative real-time RT-PCR. Furthermore, overexpression of exogenous NICE-3 by recombinant plasmids could significantly promote cell proliferation, colony formation and soft agar colony formation in Focus and WRL-68 HCC cell lines. Reversely, NICE-3 silencing by RNA interference could markedly inhibit these malignant phenotypes in YY-8103 and MHCC-97H cells. Moreover, cell cycle analysis of MHCC-97H transfected with siRNA by flow cytometry showed that NICE-3 knockdown may inhibit cell growth via arrest in G0/G1 phase and hindering entry of cells into S phase. All data of our findings indicate that NICE-3 may contribute to human hepatocellular carcinoma by promoting cell proliferation.

Decreased selenium-binding protein 1 enhances glutathione peroxidase 1 activity and downregulates HIF-1α to promote hepatocellular carcinoma invasiveness.

PURPOSE: We aimed to characterize the role of selenium-binding protein 1 (SBP1) in hepatocellular carcinoma (HCC) invasiveness and underlying clinical significance. EXPERIMENTAL DESIGN: SBP1 expression was measured in stepwise metastatic HCC cell lines by Western blotting. The role of SBP1 in HCC was investigated using siRNA. Immunofluorescence analyses were used to detect the interaction between SBP1 and glutathione peroxidase 1 (GPX1). Nineteen fresh tumor tissues and 323 paraffin-embedded samples were used to validate in vitro findings and to detect the prognostic significance of SBP1, respectively. RESULTS: Inhibition of SBP1 effectively increased cell motility, promoted cell proliferation, and inhibited apoptosis only under oxidative stress; it also greatly enhanced GPX1 activity without altering GPX1 expression and downregulated hypoxia-inducible factor-1α (HIF-1α) expression. SBP1 and GPX1 formed nuclear bodies and colocalized under oxidative stress. In freshly isolated clinical HCC tissues, decreased SBP1 was linked with increased GPX1 activity and correlated with vascular invasion. Tumor tissue microarrays indicated that SBP1 was an independent risk factor for overall survival and disease recurrence; patients with lower SBP1 expression experienced shorter overall survival periods and higher rates of disease recurrence (P < 0.001). Further analyses indicated that the predictive power of SBP1 was more significant for patients beyond the Milan criteria than patients within the Milan criteria. CONCLUSIONS: Decreased expression of SBP1 could promote tumor invasiveness by increasing GPX1 activity and diminishing HIF-1α expression in HCC; SBP1 could be a novel biomarker for predicting prognosis and guiding personalized therapeutic strategies, especially in patients with advanced HCC.

Proteasome inhibitor interacts synergistically with autophagy inhibitor to suppress proliferation and induce apoptosis in hepatocellular carcinoma.

BACKGROUND: The ubiquitin-proteasome system and autophagy-lysosome system are 2 major protein degradation pathways in eukaryotic cells, which are tightly linked to cancer. Proteasome inhibitors have been approved in clinical use against hematologic malignancies, but their application in solid tumors is uncertain. Moreover, the role of autophagy after proteasome inhibition is controversial. METHODS: Two proteasome inhibitors, 2 autophagy inhibitors, and 3 hepatocellular carcinoma (HCC) cell lines were investigated in the current study. In vitro, cell proliferation was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, cell apoptosis was evaluated by flow cytometry analysis of annexin-V/propidium iodide staining, and autophagy was evaluated by green fluorescent protein-light chain 3 (GFP-LC3) redistribution and LC3 Western blot analysis. In vivo, Ki-67 staining was used to detect cell proliferation, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining was used to detect apoptosis, and electron microscopy and p62 immunohistochemical staining were used to detect autophagy. RESULTS: Proteasome inhibitors suppressed proliferation, induced apoptosis, and activated autophagy in HCC cell lines in vitro, and autophagy exerted a protective role after proteasome inhibition. In vivo, anticancer effects of bortezomib on the MHCC-97H orthotopic model (human HCC cells) were different from the effects observed on the Huh-7 subcutaneous model (human HCC cells). The autophagy inhibitor chloroquine interacted synergistically with bortezomib to suppress proliferation and induce apoptosis in both tumor models. CONCLUSIONS: The current results indicated that simultaneous targeting of the proteasome and autophagy pathways may represent a promising method for HCC treatment.

Autophagy activation in hepatocellular carcinoma contributes to the tolerance of oxaliplatin via reactive oxygen species modulation.

PURPOSE: Understanding the roles of mammalian autophagy in cancer highlights recent advances in the pharmacologic manipulation of autophagic pathways as a therapeutic strategy for cancer. However, autophagy status and corresponding functions in hepatocellular carcinoma (HCC) after therapeutic stress remain to be clarified. This study was to determine whether the autophagic machinery could be activated after chemotherapy and the contribution of autophagy to tolerance of oxaliplatin in HCC. EXPERIMENTAL DESIGN: Autophagy activation and cell death induced by oxaliplatin were examined in two HCC cell lines as well as in vivo using an HCC model in nude mice. HCC tissue samples with or without locoregional chemotherapy before surgery were also examined by immunohistochemical and electron microscopic analysis. RESULTS: Autophagy was functionally activated in HCC cell lines and xenografts after oxaliplatin treatment. Suppression of autophagy using either pharmacologic inhibitors or RNA interference of essential autophagy gene enhanced cell death induced by oxaliplatin in HCC cells. Generation of reactive oxygen species has an important role in the induction of cell death by oxaliplatin in combination with autophagy inhibitors. Critically, the combination of oxaliplatin with autophagy inhibitor chloroquine resulted in a more pronounced tumor suppression in HCC xenografts. Furthermore, autophagy-specific protein LC3 and autophagic autophagosome formation were induced to a significantly higher level in HCC specimens that had been subjected to locoregional chemotherapy. CONCLUSIONS: Autophagy activation under therapy stress contributes to HCC tumor cell survival. Targeting the autophagy pathway is a promising therapeutic strategy to enhance the effects of chemotherapy and improve clinical outcomes in HCC patients.

Targeting autophagy enhances sorafenib lethality for hepatocellular carcinoma via ER stress-related apoptosis.

Sorafenib, a potent multikinase inhibitor, has been recognized as the standard systemic treatment for patients with advanced hepatocellular carcinoma (HCC). However, the direct functional mechanism of tumor lethality mediated by sorafenib remains to be fully characterized, and the precise mechanisms of drug resistance are largely unknown. Here, we showed sorafenib induced both apoptosis and autophagy in human HCC cells through a mechanism that involved endoplasmic reticulum (ER) stress and was independent of the MEK1/2-ERK1/2 pathway. Upregulation of IRE1 signals from sorafenib-induced ER stress was critical for the induction of autophagy. Moreover, autophagy activation alleviated the ER stress-induced cell death. Inhibition of autophagy using either pharmacological inhibitors or essential autophagy gene knockdown enhanced cell death in sorafenib treated HCC cell lines. Critically, the combination of sorafenib with the autophagy inhibitor chloroquine produced more pronounced tumor suppression in HCC both in vivo and in vitro. These findings indicated that both ER stress and autophagy were involved in the cell death evoked by sorafenib in HCC cells. The combination of autophagy modulation and molecular targeted therapy is a promising therapeutic strategy in treatment of HCC.