Production of Polyhydroxyalkanoates in Unsterilized Hyper-Saline Medium by Halophiles Using Waste Silkworm Excrement as Carbon Source.

The chlorophyll ethanol-extracted silkworm excrement was hardly biologically reused or fermented by most microorganisms. However, partial extremely environmental halophiles were reported to be able to utilize a variety of inexpensive carbon sources to accumulate polyhydroxyalkanoates. In this study, by using the nile red staining and gas chromatography assays, two endogenous haloarchaea strains: Haloarcula hispanica A85 and Natrinema altunense A112 of silkworm excrement were shown to accumulate poly(3-hydroxybutyrate) up to 0.23 g/L and 0.08 g/L, respectively, when using the silkworm excrement as the sole carbon source. The PHA production of two haloarchaea showed no significant decreases in the silkworm excrement medium without being sterilized compared to that of the sterilized medium. Meanwhile, the CFU experiments revealed that there were more than 60% target PHAs producing haloarchaea cells at the time of the highest PHAs production, and the addition of 0.5% glucose into the open fermentation medium can largely increase both the ratio of target haloarchaea cells (to nearly 100%) and the production of PHAs. In conclusion, our study demonstrated the feasibility of using endogenous haloarchaea to utilize waste silkworm excrement, effectively. The introduce of halophiles could provide a potential way for open fermentation to further lower the cost of the production of PHAs.

Urethane tetrathiafulvalene derivatives: synthesis, self-assembly and electrochemical properities.

This paper reports the self-assembly of two new tetrathiafulvalene (TTF) derivatives that contain one or two urethane groups. The formation of nanoribbons was evidenced by scanning electron microscopy (SEM) and X-ray diffraction (XRD), which showed that the self-assembly ability of T 1 was better than that of T 2 . The results revealed that more urethane groups in a molecule did not necessarily instigate self-assembly. UV-vis and FTIR spectra were measured to explore noncovalent interactions. The driving forces for self-assembly of TTF derivatives were mainly hydrogen bond interactions and π-π stacking interactions. The electronic conductivity of the T 1 and T 2 films was tested by a four-probe method.

An Andrias davidianus derived composite hydrogel with enhanced antibacterial and bone repair properties for osteomyelitis treatment.

Effective antibacterial therapy while accelerating the repair of bone defects is crucial for the treatment of osteomyelitis. Inspired by the protective mechanism of Andrias davidianus, we constructed an antibacterial hydrogel scaffold with excellent rigidity and long-term slow-release activity. While retaining the toughness of the skin secretion of Andrias davidianus (SSAD), the rigidity of the hydrogel material is increased by incorporating hydroxyapatite to meet the demands of bone-defect-filling materials. It also exerted antibacterial effects via the slow-release of vancomycin from local osteomyelitis lesions. Notably, the hydrogel can also carry a high stable recombinant miR-214-3p inhibitor (MSA-anti214). By the delivery of nano vector polyvinylamine, the long-term slow-release of MSA-anti214 is achieved to promote bone repair, making this composite hydrogel a potential SSAD-based osteomyelitis alleviator (SOA). In vitro and vivo results verified that the SOA effectively eliminated Staphylococcus aureus and repaired bone defects, ultimately mitigating the progression of osteomyelitis. This composite hydrogel extends the economic application prospects of A. davidianus and has provided new insights for the treatment of osteomyelitis. The study also explored new insights for the bone filling materials of bone defection and other skeletal system diseases.

Correlation between fungi and volatile compounds during different fermentation modes at the industrial scale of Merlot wines.

Elucidation of the relationship between fungal community development and dynamic changes in volatile components during fermentation is of great significance in controlling wine production. However, such studies on an industrial scale are rarely reported. In this study, fungal community succession during spontaneous fermentation (SPF) and inoculation fermentation (INF) of Merlot wine was monitored by a research strategy combining culture-dependent and culture-independent methods. The volatile compounds were monitored during SPF and INF by headspace solid-phase micro-extraction coupled with gas chromatography-mass spectrometry technology. The Spearman correlation coefficient was also used to investigate the interplay between fungal communities and volatile compounds. We found that fungal community diversity in SPF decreased as fermentation progressed but was significantly higher than that of INF. Starmerella and Kazachstania were the dominant non-Saccharomyces genera in Merlot wine during SPF. However, the presence of commercial yeasts and sulphur dioxide led to a sharp decrease or the disappearance of non-Saccharomyces genera during INF. Spearman correlation analysis revealed that all major volatiles were positively correlated with most functional microbiotas except P. fermentans, S. bacillaris, E. necator, and D. exigua in INF. In SPF, most non-Saccharomyces were negatively correlated with core volatiles, whereas K. humilis, M. laxa, P. kluyveri, and A. japonicus were positively correlated with the major volatiles, especially some higher alcohols (isopentol, heptanol) and terpenes (linalool, citronellol). S. cerevisiae was positively correlated with most of the main volatile substances except ethyl isovalerate and isoamyl acetate. These findings provide a reference for comprehending the diverse fermentation methods employed in the wine industry and improving the quality of Merlot wines.

Dynamic analysis of microbial communities and flavor properties in Merlot wines produced from inoculation and spontaneous fermentation.

The microbiota is of great importance in forming flavor compounds and improving sensory characteristics during wine fermentation. Understanding microbial succession is critical for controlling its contribution to wine flavor with predictable sensory quality. In this study, microbial community composition and characteristic flavor compounds were identified during the inoculation fermentation (IF) and spontaneous fermentation (SF) to provide a basis for exploring the relationship between these microorganisms and volatile components. The results demonstrated that SF had higher fungal community diversity and lower bacterial community diversity than IF. Eleven (11) fungal and 10 bacterial genera (relative abundance > 0.1 %) were considered beneficial microbiota. Saccharomyces, Hanseniaspora, and Alternaria were the leading fungal genera in SF. Massilia, Nesterenkonia, and Halomonas were the predominant bacteria in IF, while Tatumella and Ochrobactrum were mainly from SF. In addition, the microbial community composition was reshaped via correlational analysis between microbiota succession and physicochemical properties, mainly attributed to the changes in environmental factors during fermentation. The SF wines had more aromatic higher alcohols, acetate esters, and terpenes. Also, the sensory evaluation showed that the SF wines were characterized by more fruity, floral, intense, and typical aromas. The associations between the microbial community and the volatile components indicated that the dominant species largely determined the characteristic flavor compounds during fermentation.

Study on the inhibitory effect of selective estrogen receptor modulators on Streptococcus mutans.

OBJECTIVES: To screen small-molecule antibacterial drugs and investigate the antibacterial effect and mechanism of selective estrogen receptor modulators (SERMs) against Streptococcus mutans (S.mutans). METHODS: The minimum inhibitory concentration of 426 Food and Drug Administration (FDA)-approved small-molecule drugs against S. mutans was determined using the microdilution method, and the target of SERMs acting on S. mutans was explored by employing a random transposon mutant library. RESULTS: Among the 426 FDA-approved SERMs, toremiphene, tamoxifen, clomiphene, and raloxifene exhibited excellent antibacterial effects against S.mutans. Results of mutant library screening showed that the two mutant strains were resistant to clomiphene. The gene sequence of the resistant strains showed that the transposon insertion sites were located in the genes of smu_546 and smu_874. CONCLUSIONS: SERMs, such as toremifene, tamoxifen, clomiphene, and raloxifene, exerted obvious antibacterial effects on S. mutans, and their targets may be proteins expressed by smu_546 and smu_874 gene.

Effects of hydrodynamic conditions on the composition, spatiotemporal distribution of different extracellular polymeric substances and the architecture of biofilms.

Microbial biofilms are common on abiotic and biotic surfaces, especially in rivers, which drive crucial ecosystem processes. The microorganisms of biofilms are surrounded by a self-produced extracellular polymeric substance (EPS). In this study, we investigated the effects of different hydrodynamic conditions on the composition, spatiotemporal distribution of different extracellular polymeric substances, and the architecture of biofilms. Multidisciplinary methods offer complementary insights into complex architecture correlations in biofilms. The biofilms formed in turbulent flow with high shear force were thin but dense. However, the biofilms formed under laminar flow conditions were thick but relatively loose. The thickness and compactness of the biofilms formed in the transitional flow were different from those of the other biofilms. The compact structure of the biofilm helped to resist shear forces to minimize detachment. Under the turbulent flow condition, bacteria, exopolysaccharides, and extracellular proteins permeated through the biofilm, and more extracellular polysaccharides enveloped bacteria and extracellular proteins. However, under the transitional flow condition, the extracellular polysaccharides and proteins were fewer than those under the turbulent flow condition; bacteria and algae were seen more prominently in the upper layer of the biofilm. Under the laminar flow condition, the distribution of extracellular polysaccharides, extracellular proteins, and bacteria was relatively uniform throughout the biofilm. The number of extracellular polysaccharides was greater than that of extracellular proteins. The total number of EPS in the biofilm was the largest under turbulent flow condition, followed by that under transitional flow condition and then under laminar flow condition. This study also observed that soluble EPS (S-EPS) were secreted first, followed by loosely bound EPS (LB-EPS) and tightly bound EPS (TB-EPS). In particular, the adhesion of LB-EPS and flocculation capability of TB-EPS play some role in regulating biofilm formation. This study would help to perfect the five-stages theory of biofilm formation.

Study on the production of high 3HV content PHBV via an open fermentation with waste silkworm excrement as the carbon source by the haloarchaeon Haloferax mediterranei.

Silkworm excrement is hard to be degraded or bio-utilized by environmental microorganisms due to its high content of heavy metals and antimicrobial biomacromolecules in mulberry leaves. In traditional Chinese silk industry, the silkworm excrement results in environmental problems. In this study, the silkworm excrement after chlorophyll ethanol-extraction was researched. An open fermentation strategy was developed using the silkworm excrement as the sole or partial carbon source by haloarchaea to accumulate polyhydroxyalkanoates. As a haloarchaeon with strong carbon source utilization ability, Haloferax mediterranei was found to accumulate a certain amount of poly(3-hydroxybutyrate-co-3-hydroxyvalerate; PHBV) using waste silkworm excrement. The results showed that the addition of silkworm excrement into glucose based fermentation medium can significantly improve the production of PHBV. Using a mixture carbon source including the extract of silkworm excrement and glucose (with a 1:1 carbon content ratio), the yield of PHBV was 1.73 ± 0.12 g/l, which showed a 26% increase than that of fermentation without the silkworm excrement addition. When the NaCl content of medium was set to approximately 15%, fermentation without sterilization was performed using silkworm excrement as the carbon source. Moreover, the addition of the silkworm excrement extract could increase the 3-hydroxyvalerate (3 HV) content of PHBV regardless of the sterilization or non-sterilization fermentation conditions. When using silkworm excrement as the sole carbon source, the 3 HV content was as high as 16.37 ± 0.54 mol %. The real-time quantitative PCR results showed that the addition of the silkworm excrement could specifically enhance the expression of genes involved in the aspartate/2-ketobutyric acid pathway related to 3 HV synthesis in H. mediterranei, and further analysis of the amino acid of the silkworm excrement suggested that the high content of threonine in the silkworm excrement might be the reason for the increase of 3 HV content. Taken together, the success of non-sterile fermentation in hypersaline condition using haloarchaea implied a novel way to reuse the silkworm excrement, which not only reduces the production costs of PHBV, but also is conducive to environmental protection.

Classical Dichotomy of Macrophages and Alternative Activation Models Proposed with Technological Progress.

Macrophages are important immune cells that participate in the regulation of inflammation in implant dentistry, and their activation/polarization state is considered to be the basis for their functions. The classic dichotomy activation model is commonly accepted, however, due to the discovery of macrophage heterogeneity and more functional and iconic exploration at different technologies; some studies have discovered the shortcomings of the dichotomy model and have put forward the concept of alternative activation models through the application of advanced technologies such as cytometry by time-of-flight (CyTOF), single-cell RNA-seq (scRNA-seq), and hyperspectral image (HSI). These alternative models have great potential to help macrophages divide phenotypes and functional genes.

Prognostic Value of Inflammatory and Tumour Markers in Small-Duct Subtype Intrahepatic Cholangiocarcinoma after Curative-Intent Resection.

Intrahepatic cholangiocarcinoma (ICC) is characterised by heterogeneity, and it can be subdivided into small-duct and large-duct types. Inflammatory and tumour markers could effectively predict prognosis in many cancers, but no similar studies have been conducted in the histological subtypes of ICC. A total of 102 and 72 patients with ICC undergoing curative-intent resection were retrospectively subclassified into large-duct and small-duct types by chemical staining, respectively. The prognostic value of inflammatory and tumour markers was studied for the first time in histological subtypes of ICC by using a Cox regression model. A novel predictor named prognostic inflammatory index (PII) was proposed and defined as neutrophil × monocyte/lymphocyte count (109/L). Survival analysis showed that PII, neutrophil-to-lymphocyte ratio (NLR), lymphocyte-to-monocyte ratio (LMR), carcinoembryonic antigen (CEA), carbohydrate antigen 19-9 (CA19-9), CA242, and ferritin were all predictors of DFS and OS in patients with ICC (P < 0.040). Subgroup analysis showed that PII, CA19-9, and ferritin were risk predictors of disease-free survival (DFS) and overall survival (OS) in small-duct type ICC (P < 0.015). In addition, in small-duct type ICC, NLR and LMR were correlated with OS (P < 0.025), whilst CEA and CA242 were correlated with DFS (P ≤ 0.010). In conclusion, PII is a convenient and efficient inflammatory predictor of DFS and OS in ICCs and their small-duct type. NLR and LMR, rather than platelet-to-lymphocyte ratio, were correlated with OS in small-duct type ICC. In addition, ferritin may be a supplement to CA19-9 in stratifying the survival outcome of patients with small-duct type ICC.

Analyses of production capacity of 89Sr and 90Sr in the 2 MW molten salt reactor.

The production capacity of 89Sr and 90Sr in the 2 MW MSR are evaluated. The gaseous 89Kr and 90Kr are extracted from the core through the helium bubbling system, and then decay to 89Sr and 90Sr, respectively. In order to improve purity of 89Sr product, two cooling devices are adopted in the 89Sr and 90Sr production system. The annual yields of 89Sr and 90Sr are about 9000 Ci and 32 Ci, respectively, and the impurity of 89Sr product is less than 2 ppm which can meet the medical requirement.

Identification of potential biomarkers in dengue via integrated bioinformatic analysis.

Dengue fever virus (DENV) is a global health threat that is becoming increasingly critical. However, the pathogenesis of dengue has not yet been fully elucidated. In this study, we employed bioinformatics analysis to identify potential biomarkers related to dengue fever and clarify their underlying mechanisms. The results showed that there were 668, 1901, and 8283 differentially expressed genes between the dengue-infected samples and normal samples in the GSE28405, GSE38246, and GSE51808 datasets, respectively. Through overlapping, a total of 69 differentially expressed genes (DEGs) were identified, of which 51 were upregulated and 18 were downregulated. We identified twelve hub genes, including MX1, IFI44L, IFI44, IFI27, ISG15, STAT1, IFI35, OAS3, OAS2, OAS1, IFI6, and USP18. Except for IFI44 and STAT1, the others were statistically significant after validation. We predicted the related microRNAs (miRNAs) of these 12 target genes through the database miRTarBase, and finally obtained one important miRNA: has-mir-146a-5p. In addition, gene ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment were carried out, and a protein-protein interaction (PPI) network was constructed to gain insight into the actions of DEGs. In conclusion, our study displayed the effectiveness of bioinformatics analysis methods in screening potential pathogenic genes in dengue fever and their underlying mechanisms. Further, we successfully predicted IFI44L and IFI6, as potential biomarkers with DENV infection, providing promising targets for the treatment of dengue fever to a certain extent.

Dimethylation of eEF1A at Lysine 55 Plays a Key Role in the Regulation of eEF1A2 on Malignant Cell Functions of Acute Myeloid Leukemia.

OBJECTIVE: This study aimed to explore whether eukaryotic translation elongation factor 1 alpha 2 affected cell proliferation, migration, and apoptosis via regulating the dimethylation of eukaryotic translation elongation factor 1 alpha at lysine 55 in acute myeloid leukemia. METHODS: The expressions of eukaryotic translation elongation factor 1 alpha 2 and dimethylation of eukaryotic translation elongation factor 1 alpha at lysine 55 in acute myeloid leukemia cell lines and human normal bone marrow mononuclear cells (as control) were assessed. Control CRISPR-Cas9 lentivirus, eukaryotic translation elongation factor 1 alpha 2 knockout CRISPR-Cas9 lentivirus, vector plasmid, eukaryotic translation elongation factor 1 alpha 2 wild type overexpression plasmid, and eukaryotic translation elongation factor 1 alpha 2 with a K55R substitution overexpression plasmid were transfected into AML-193 and Kasumi-1 cells combined or alone, and were accordingly divided into 4 groups (Sgcontrol + vector group, SgeEF1A2 + vector group, SgeEF1A2 + eEF1A2WT group, and SgeEFIA2 + eEF1A2K55R group). RESULTS: Eukaryotic translation elongation factor 1 alpha 2 and dimethylation of eukaryotic translation elongation factor 1 alpha at lysine 55 expressions were higher in AML-193, Kasumi-1, and KG-1 cell lines compared to the control. In AML-193 and Kasumi-1 cells, the knockout and compensated experiments revealed that eukaryotic translation elongation factor 1 alpha 2 promoted cell proliferation and migration but repressed apoptosis. Additionally, the knockout of eukaryotic translation elongation factor 1 alpha 2 decreased dimethylation of eukaryotic translation elongation factor 1 alpha at lysine 55 expression, meanwhile, eukaryotic translation elongation factor 1 alpha 2 wild type overexpression enhanced while eukaryotic translation elongation factor 1 alpha 2 with a K55R substitution overexpression did not influence the dimethylation of eukaryotic translation elongation factor 1 alpha at lysine 55 expression. Furthermore, eukaryotic translation elongation factor 1 alpha 2 wild type overexpression promoted cell proliferation, enhanced migration, and decreased apoptosis, but eukaryotic translation elongation factor 1 alpha 2 with a K55R substitution overexpression did not influence these cellular functions in AML-193 and Kasumi-1 cells, suggesting the implication of dimethylation of eukaryotic translation elongation factor 1 alpha at lysine 55 in eukaryotic translation elongation factor 1 alpha 2 mediated oncogenesis of acute myeloid leukemia. CONCLUSION: Eukaryotic translation elongation factor 1 alpha 2 and its dimethylated product may serve as therapeutic targets, and these findings may provide support for exploring novel strategies in acute myeloid leukemia treatment.

Directed C3-Alkoxymethylation of Indole via Three-Component Cascade Reaction.

An efficient and regioselective C3-alkoxymethylation of indoles has been developed with aldehydes and alcohols via three-component cascade reaction under transition-metal free conditions. This method allows for rapid access to a variety of C3-alkoxymethylaed free (N-H) indole in up to 98% yield with excellent regioselectivity. The titled products are useful building blocks in organic synthesis.

High GOLPH3 expression is associated with poor prognosis and invasion of hepatocellular carcinoma.

Golgi phosphoprotein 3 (GOLPH3) overexpression has previously been associated with the progression of several solid tumors, which resulted in adverse clinical outcomes. The present study aimed to determine the expression and prognostic significance of GOLPH3 in human hepatocellular carcinoma (HCC). GOLPH3 expression was examined using western blot analysis of 30 paired samples of HCC and adjacent non­cancerous liver tissues. GOLPH3 expression levels were also assessed using immunohistochemistry in 180 HCC samples and paired controls. In addition, the association of GOLPH3 expression with clinicopathological features and clinical outcome was analyzed. Furthermore, the effect of GOLPH3 on HCC cell proliferation and invasion were determined. Western blot analysis revealed that GOLPH3 expression was significantly elevated in HCC tissue compared with that of the matched adjacent non­cancerous liver tissue. In addition, the results of the immunohistochemical analysis demonstrated that GOLPH3 expression was positively correlated with the Edmondson­Steiner grade (P=0.006), vascular invasion (P=0.002) and serum α feto­protein levels (P=0.015). GOLPH3 expression was found to be an independent factor for predicting the poor overall survival of HCC patients (hazard ratio, 2.01; 95% confidence interval, 1.26­3.64; P=0.025). In addition, GOLPH3 silencing inhibited the proliferation, invasion and migration of HCC cell lines in vitro. In conclusion, the results of the present study demonstrated that high GOLPH3 levels may be a potential biomarker for the poor prognosis of patients with HCC.

Epigallocatechin-3-gallate regulates the expression of Kruppel-like factor 4 through myocyte enhancer factor 2A.

Epigallocatechin-3-gallate (EGCG), a powerful antioxidant and free ion scavenger found in green tea, exhibits inhibitory effects on different stages of tumorigenesis. Within gastric cancer cells, the transcription factor Kruppel-like factor 4 (KLF4) is downregulated, and it is possible that EGCG exerts its anti-tumorigenic function through modulation of KLF4 expression. In order to examine the effects of EGCG on KLF4 in a gastric tumor model, we treated the gastric cancer cell line NCI-N87 with EGCG. We found that EGCG treatment results in increased expression of KLF4 and alters expression of the KLF4 target genes p21, CDK4, and cyclin D1. EGCG inhibits the growth of NCI-N87 cells in a time- and dose-dependent manner through arresting the cell cycle in the G0/G1 phase. Furthermore, terminal deoxynucleotidyl transferase dUTP nick end labeling assay and 4',6-diamidino-2-phenylindole staining revealed that EGCG is able to promote apoptosis of NCI-N87 cells. The suppressive effects of EGCG on cell growth and cell cycle protein expression are eliminated by decreasing KLF4 mRNA using siRNA and are magnified by overexpressing KLF4. Using KLF4 reporter constructs, we verified that the elevated expression induced by EGCG was mediated by increasing levels of activated MEF2A, which bound to the promoter region of KLF4. Taken together, this is the first time that EGCG is reported to increase the expression of KLF4, suggesting a novel mechanisms in gastric cancer treatment.