Functional characterization of a novel flavin reductase from a deep-sea sediment metagenomic library and its application for indirubin production.

Microbial synthesis is a desirable approach to produce indirubin but suffers from low synthetic efficiency. Insufficient supply of reduced flavins is one major factor limiting synthetic efficiency. To address this, a novel flavin reductase, MoxB, was discovered through screening of the metagenomic library. MoxB showed a strong preference for NADH over NADPH as the electron source for FMN/FAD reduction and exhibited the highest activity at pH 8.0 and 30°C. It displayed remarkable thermostability by maintaining 80% of full activity after incubation at 60°C for 1 h. Furthermore, MoxB showed great organic solvent tolerance and its activity could be significantly increased by bivalent metal ions. In addition, heterologous expression of the moxB gene in the indirubin-producing E. coli significantly improved indirubin production up to 15.12-fold. This discovery expands the understanding of flavin reductases and provides a promising catalytic tool for microbial indirubin production.IMPORTANCEMuch effort has been exerted to produce indirubin using engineered Escherichia coli, but high-level production has not been achieved so far. Insufficient supply of reduced flavins is one key factor limiting the catalytic efficiency. However, the flavin reductases involved in indirubin biosynthesis have not been hitherto reported. Discovery of the novel flavin reductase MoxB provides a useful tool for enhancing indirubin production by E. coli. Overexpression of MoxB in indirubin-producing E. coli increased indirubin production by 15.12-fold in comparison to the control strain. Our results document the function of flavin reductase that reduces flavins during indirubin biosynthesis and provide an important foundation for using the flavin reductases to improve indirubin production by engineered microorganisms.

Metal-free synthesis of 1,3-dichloro-1,5-diarylpentan-5-ones via cascade oxidative radical addition of styrenes with CHCl3.

A novel and efficient metal-free cascade oxidative radical addition of styrenes is developed for the construction of 1,3-dichloro-1,5-diarylpentan-5-ones. This protocol presents a practical one-pot procedure that delivers highly functionalized 1,3-dichloro-1,5-diarylpentan-5-ones in moderate-to-good yields with a broad substrate scope under mild conditions.

Characterization of a novel monooxygenase originating from a deep-sea sediment metagenomic library.

Oxygenases are important biocatalysts to produce many industrially important biomolecules. Here, a novel oxygenase, named MoxA, was identified through screening of a deep-sea sediment metagenomic library. Sequence analysis showed MoxA contains 424 amino acid residues with a predicated molecular mass of 46.9 kDa. Multiple sequence alignment and phylogenetic analysis indicated the sequence might be a new member of monooxygenase subfamily. A recombinant MoxA was obtained through the functional expression of moxA gene in Escherichia coli. Characterization of the purified MoxA indicated that it is an alkaline oxygenase showing maximal activity at pH 8.0. The optimal temperature of MoxA was 37 ℃, and it retained more than 70% of its initial activity after 1 h at 20-50 ℃ exhibiting good thermostability. Furthermore, effect of metal ions and organic solvents on enzymatic activity was investigated, and the results showed that the activity of MoxA was enhanced by Cu2+, Zn2+, Co2+ and Mg2+ at 1 mM, and by Co2+, Ca2+ and Mg2+ at 5 mM. Moreover, the recombinant strain harboring MoxA was used as a whole-cell biocatalyst for the efficient biosynthesis of indigo showing promising conversion efficiency. The biochemical properties of MoxA indicated that it would provide great contribution for the indigo bioproduction. KEY POINTS: • A novel monooxygenase from a metagenomic library was characterized. • The activity of MoxA was enhanced by metal ions at 1 mM and 5 mM. • MoxA has an optimal temperature of 37 ℃ and exhibited high conversion capacity.

You are how you speak: The roles of vocal pitch and semantic cues in shaping social perceptions.

Social perceptions of speakers are influenced by their voice information, including vocal characteristics and semantic content. Our study investigated how individuals' warmth- and competence-related perceptions of speakers were affected by vocal pitch levels (i.e., high/low) and three kinds of semantic cues (i.e., prosocial, antisocial, and neutral) simultaneously. We have three key findings. First, antisocial cues negatively affected social perceptions, regardless of speakers' gender. However, prosocial cues did not have positive impacts on evaluations of speakers because ratings were similar between prosocial cues and neutral cues. Second, female vocal pitch mattered for warmth-related perceptions but not for competence-related perceptions. The role of semantic cues should be additionally considered when investigating the impact of male vocal pitch on these perceptions. For example, higher-pitched men in prosocial contexts were perceived as warmer, while low-pitched men in antisocial contexts were judged as more competent. Third, the connection between vocal pitch and two kinds of perceptions showed an opposite trend, in which high pitch was related to more warmth but less competence, while the low pitch was associated with less warmth but more competence. These findings extend the understanding of the role of vocal pitch in the formation of stereotypes of strangers in different semantic contexts.

Catalpol induces apoptosis in breast cancer in vitro and in vivo: Involvement of mitochondria apoptosis pathway and post-translational modifications.

Breast cancer is a fatal cancer with the highest mortality in female. New strategies for anti-breast cancer are still urgently needed. Catalpol, an iridoid glycoside extracted from the traditional Chinese medicinal plant Rehmannia glutinosa, has shown anticancer efficacy in various cancer cells. However, its effect on breast cancer remains unclear. In this study, we aim to investigate the anti-breast cancer activity of catalpol and elucidate its underlying mechanism. Cell counting kit-8 (CCK-8) and morphology change showed that catalpol could inhibit the proliferation and viability of MCF-7 cells. Catalpol administration reduced the tumor volume in xenograft model. Catalpol induced apoptosis in MCF-7 cells confirmed by Hoechst 33342 staining and Annexin V-FITC/PI double staining. In vivo, catalpol also induced apoptosis as seen from the increased level of terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) in tumor. According to JC-1 and Dichlorodi-hydrofluorescein Diacetate (DCFH-DA) staining, loss of mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) generation was found in MCF-7 cells treated with catalpol. Furthermore, catalpol also increased the level of cytoplasmic cytochrome c and activity of caspase-3 in MCF-7 cells. Likewise, histopathological and immunohistochemical (IHC) assay also found that catalpol enhanced the levels of cytochrome c and caspase-3 in breast cancer tissues. Ultimately, acetylation, 2-hydroxyisobutyrylation and lactylation were dramatically increased, whereas succinylation, malonylation and phosphorylation were markedly decreased in the breast cancer tumor treated with catalpol. Taken together, catalpol inhibited breast cancer in vitro and in vivo through induction of apoptosis via mitochondria apoptosis pathway and regulation of protein post-translational modifications (PTMs). Thus, it can be considered as an excellent candidate compound for treatment of breast cancer.

Schisandra chinensis essential oil attenuates acetaminophen-induced liver injury through alleviating oxidative stress and activating autophagy.

CONTEXT: Schisandra chinensis (Turcz.) Baill. (Magnoliaceae) essential oil (SCEO) composition is rich in lignans that are believed to perform protective effects in the liver. OBJECTIVE: This study investigates the effects of SCEO in the treatment of acetaminophen (APAP)-induced liver injury in mice. MATERIALS AND METHODS: C57BL/6 mice (n = 56) were randomly divided into seven groups: normal; APAP (300 mg/kg); APAP plus bicyclol (200 mg/kg); APAP plus SCEO (0.25, 0.5, 1, 2 g/kg). Serum biochemical parameters for liver function, inflammatory factors, and antioxidant activities were determined. The protein expression levels of Nrf2, GCLC, GCLM, HO-1, p62, and LC3 were assessed by western blotting. Nrf2, GCLC, HO-1, p62, and LC3 mRNA were detected by real-time PCR. RESULTS: Compared to APAP overdose, SCEO (2 g/kg) pre-treatment reduced the serum levels of AST (79.4%), ALT (84.6%), TNF-α (57.3%), and IL-6 (53.0%). In addition, SCEO (2 g/kg) markedly suppressed cytochrome P450 2E1 (CYP2E1) (15.4%) and attenuated the exhaustion of GSH (43.6%) and SOD (16.8%), and the accumulation of MDA (22.6%) in the liver, to inhibit the occurrence of oxidative stress. Moreover, hepatic tissues from our experiment revealed that SCEO pre-treatment mitigated liver injury caused by oxidative stress by increasing Nrf2, HO-1, and GCL. Additionally, SCEO activated autophagy, which upregulated hepatic LC3-II and decreased p62 in APAP overdose mice (p < 0.05). DISCUSSION AND CONCLUSIONS: Our evidence demonstrated that SCEO protects hepatocytes from APAP-induced liver injury in vivo and the findings will provide a reliable theoretical basis for developing novel therapeutics.

SLC45A4 promotes glycolysis and prevents AMPK/ULK1-induced autophagy in TP53 mutant pancreatic ductal adenocarcinoma.

BACKGROUND: Somatic mutations of the TP53 gene occur frequently in pancreatic ductal adenocarcinoma (PDA). Solute carrier family 45 member A4 (SLC45A4) is a H+ -dependent sugar cotransporter. The role of SLC45A4 in PDA, especially in TP53 mutant PDA, remains poorly understood. METHODS: We explored the TCGA datasets to identify oncogenes in TP53 mutant PDA. MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium], colony formation and 5-ethynyl-2'-deoxyuridine (Edu) assays were performed to investigate the function of SLC45A4 in vitro. Glucose consumption, lactate production and ATP production were detected to evaluate glucose utilization. Extracellular acidification rate and oxygen consumption rate assays were used to evaluate glycolysis and oxidative phosphorylation. The subcutaneous xenotransplantation models were conducted to explore the function of SLC45A4 in vivo. RNA-sequencing and gene set enrichment analysis were employed to explore the biological alteration caused by SLC45A4 knockdown. Western blotting was performed to evaluate the activation of glycolysis, as well as the AMPK pathway and autophagy. RESULTS: SLC45A4 was overexpressed in PDA for which the expression was significantly higher in TP53 mutant PDA than that in wild-type PDA tissues. Moreover, high level of SLC45A4 expression was tightly associated with poor clinical outcomes in PDA patients. Silencing SLC45A4 inhibited proliferation in TP53 mutant PDA cells. Knockdown of SLC45A4 reduced glucose uptake and ATP production, which led to activation of autophagy via AMPK/ULK1 pathway. Deleting SLC45A4 in TP53 mutant HPAF-II cells inhibited the growth of xenografts in nude mice. CONCLUSIONS: The present study found that SLC45A4 prevents autophagy via AMPK/ULK1 axis in TP53 mutant PDA, which may be a promising biomarker and therapeutic target in TP53 mutant PDA.

[Research progress of correlation between multidrug resistance and transporters and traditional Chinese medicine intervention].

With the emergence of drug resistance in Western medicine, the repeated administration of clinical first-line drugs becomes more severe. There are many factors leading to multidrug resistance(MDR), so it is very difficult to solve the problem. Since traditional Chinese medicine(TCM) has been used in the field of MDR in recent years, the research on the transporter-associated drug resistance and intervention of TCM has gradually become a hot spot. Therefore, in order to further explore the relationships among drug resistance, transporters, and TCM intervention, we review the relevant research progress in recent years and comb the achievements and limitations of this research at present. In the end, we put forward the research direction of changing body's ADME through the host's transporters and gastrointestinal flora, which provides new ideas for future research.

The palladium-catalyzed direct C3-cyanation of indoles using acetonitrile as the cyanide source.

The ligand-free palladium-catalyzed C3-cyanation of indoles via direct C-H functionalization was achieved. This protocol, utilizing CH3CN as a green and readily available cyanide source, produced the desired products in moderate to good yields through transition-metal-catalyzed C-CN bond cleavage.

Discovery of novel indazole-acylsulfonamide hybrids as selective Mcl-1 inhibitors.

Overexpressing myeloid cell leukemia sequence 1 (Mcl-1) protein is an important way to confer the resistance of cancer cells to conventional anti-cancer treatments. Therefore, developing Mcl-1 inhibitors has become an attractive strategy for cancer therapy. In the studies, a series of new indazole-acylsulfonamide hybrids were designed, synthesized and evaluated as potent Mcl-1 inhibitors. Among them, the most potent compound 17 (Ki = 0.43 µM) showed a little better inhibitory activity against Mcl-1 protein than positive control AT-101 (Ki = 0.45 µM). Pleasingly, it displayed > 40-fold selectivity over Bcl-2 (Ki = 18 µM) and Bcl-xL (no activity). Furthermore, compound 17 had good inhibitory activities against PC-3, MDA-MB-231 and K562 cells (IC50 = 12.3, 10.6 and 6.62 µM, respectively) and could effectively induce apoptosis and the activation of caspase-3 in a dose-dependent manner in K562 cells.

Multiuser Time-Energy Entanglement Swapping Based on Dense Wavelength Division Multiplexed and Sum-Frequency Generation.

Perfect entanglement swapping, which can be realized without the postselection by using the nonlinear optical technology, provides an important way toward generating the large-scale quantum network. We explore an entanglement-swapping-based dense wavelength division multiplexed network in the experiment. Four users receive single quantum states at different wavelengths, and we perform a time-energy entanglement swapping operation based on the sum-frequency generation to make users fully connected in the network. The results show that the fidelity of the entangled state is larger than 90% and is independent of the number of users. Our Letter demonstrates the feasibility of a proposed multiuser network, and hence paves a route toward a variety of quantum applications, including entanglement-swapping-based quantum direct communication.

Deposited Nanoparticles Can Promote Air Clogging of Piezoelectric Inkjet Printhead Nozzles.

Piezoelectric inkjet printing is susceptible to printhead clogging when printing with inks that contain dispersed particles. This paper investigates the mechanisms by which 28-530 nm nanoparticle dispersions induce printhead clogging without forming large aggregates or thick deposited layers on printhead surfaces. Printing experiments were combined with nanoparticle deposition studies and with experiments where inks were pumped through printheads at a constant flow rate with a syringe pump. Submonolayer coverages of hydrophobic cationic polystyrene nanoparticles adhering to printhead surfaces promote rapid clogging by trapped air that enters from the nozzle opening. We propose that the deposited particles distort the shape of the ink/air meniscus, possibly causing air entrainment, and promote air bubble adhesion to the interior printhead surfaces. The printer's purge-blot cleaning procedure removes air clogs, but the clogs quickly reform when printing is resumed because the adsorbed nanoparticles are not removed by the cleaning procedure. Nondepositing anionic hydrophobic nanoparticles cause much less clogging, possibly because of filtration of trace large aggregates. Colloidal stability is a necessary but not sufficient criterion for ink dispersions; the ink particles must not adsorb onto the printhead surfaces. Thus, alternate surface chemistries for the printhead and ink particle surfaces may be required to print hydrophobic ink materials.

LncRNA PVT1 triggers Cyto-protective autophagy and promotes pancreatic ductal adenocarcinoma development via the miR-20a-5p/ULK1 Axis.

BACKGROUND: Defective autophagy is thought to contribute to the pathogenesis of many diseases, including cancer. Human plasmacytoma variant translocation 1 (PVT1) is an oncogenic long non-coding RNA that has been identified as a prognostic biomarker in pancreatic ductal adenocarcinoma, but how PVT1 operates in the regulation of autophagy in pancreatic ductal adenocarcinoma (PDA) is unclear. METHODS: PVT1 expression level was detected by quantitative real-time polymerase chain reaction (qRT-PCR) and hybridization in situ (ISH). Western blot or qRT-PCR was performed to assess the ULK1 protein or mRNA level. Autophagy was explored via autophagic flux detection under a confocal microscope and autophagic vacuoles investigation under a transmission electron microscopy (TEM). The biological role of PVT1 in autophagy and PDA development was determined by gain-of-function and loss-of-function assays. RESULTS: We found that PVT1 levels paralleled those of ULK1 protein in PDA cancer tissues. PVT1 promoted cyto-protective autophagy and cell growth by targeting ULK1 both in vitro and in vivo. Moreover, high PVT1 expression was associated with poor prognosis. Furthermore, we found that PVT1 acted as sponge to regulate miR-20a-5p and thus affected ULK1 expression and the development of pancreatic ductal adenocarcinoma. CONCLUSIONS: The present study demonstrates that the "PVT1/miR-20a-5p/ULK1/autophagy" pathway modulates the development of pancreatic ductal adenocarcinoma and may be a novel target for developing therapeutic strategies for pancreatic ductal adenocarcinoma.

Multiple-DWDM-channel heralded single-photon source based on a periodically poled lithium niobate waveguide.

We report on the experimental realization of a multiple-DWDM-channel heralded single-photon source in a periodically poled lithium niobate waveguide. Our single photon at the telecom wavelength covers more than 40 channels of the ITU grid. All channels have virtually identical efficiencies, and the multi-photon emission probability is reduced by a factor up to more than 150 compared to a Poissonian light source. Together with the all-fiber structure, all these advantages make our heralded single-photon source suitable for real-world quantum networks. The implementation with a 50 MHz pulsed laser provides a data rate compatible with current quantum communication systems, while being able to be pumped at higher repetition rates.

MiR-143 Targeting TAK1 Attenuates Pancreatic Ductal Adenocarcinoma Progression via MAPK and NF-κB Pathway In Vitro.

BACKGROUND: Transforming growth factor (TGF)-ß-activated kinase 1 (TAK1) is one of the major regulators of inflammation-induced cancer cell growth and progression. MiR-143 dysregulation is a common event in a variety of human diseases including pancreatic ductal adenocarcinoma (PDA). AIMS: To identify the interaction between TAK1 and miR-143 in PDA. METHODS: Data mining of TAK1 expression in PDA patient gene profiling was conducted. QRT-PCR and western blot were performed to detect the expression of TAK1 in PDA tissues and cell lines. Ectopic miR-143 and TAK1 were introduced to PDA cells. Cell growth, apoptosis and migration were examined. Xenograft models were used to examine the function of TAK1 in vivo. Western blot and luciferase assay were carried out to investigate the direct target of miR-143. RESULTS: PDA patient gene profiling data (GSE15471 and GSE16515) showed that TAK1 mRNA was aberrantly up-regulated in PDA tissues. TAK1 protein levels were overexpressed in PDA tissues and cell lines. Overexpression of TAK1 was strongly associated with positive lymph node metastasis. Inhibition of TAK1 suppressed cell growth, migration, and induced cell apoptosis in vitro and in vivo. Further studies demonstrated that TAK1 was a direct target gene of miR-143. MiR-143 also inhibited PDA cells proliferation and migration, induced apoptosis and G1/S arrest. Moreover, TAK1 depletion inactivated MAPK and NF-κB pathway, mimicking the function of miR-143. CONCLUSIONS: The study highlights that miR-143 acts as a tumor suppressor in PDA through directly targeting TAK1, and their functional regulation may provide potential therapeutic strategies in clinics.

Use of subsequent PET/CT in diffuse large B-cell lymphoma patients in complete remission following primary therapy.

Interim 18F-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (I-PET/CT) is a powerful tool for monitoring the response to therapy in diffuse large B-cell lymphoma (DLBCL). This retrospective study aimed to determine when and how to use I-PET/CT in DLBCL. A total of 197 patients treated with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) were enrolled between October 2005 and July 2011; PET/CT was performed at the time of diagnosis (PET/CT0), after 2 and 4 cycles of chemotherapy (PET/CT2 and PET/CT4, respectively), and at the end of treatment (F-PET/CT). According to the International Harmonization Project for Response Criteria in Lymphoma, 110 patients had negative PET/CT2 scans, and 87 had positive PET/CT2 scans. The PET/CT2-negative patients had significantly higher 3-year progression-free survival rate (75.8% vs. 38.2%) and 3-year overall survival rate (93.5% vs. 55.6%) than PET/CT2-positive patients. All PET/CT2-negative patients remained negative at PET/CT4, but 3 were positive at F-PET/CT. Among the 87 PET/CT2-positive patients, 57 remained positive at F-PET/CT, and 32 progressed during chemotherapy (15 at PET/CT4 and 17 at F-PET/CT). Comparing PET/CT4 with PET/CT0, 7 patients exhibited progression, and 8 achieved partial remission. Comparing F-PET/CT with PET/CT0, 10 patients exhibited progression, and 7 achieved partial remission. In conclusion, our results indicate that I-PET/CT should be performed after 2 rather than 4 cycles of immunochemotherapy in DLBCL patients. There is a limited role for subsequent PET/CT in the detection of relapse in PET/CT2-negative patients, but repeat PET/CT is required if the PET/CT2 findings are positive.

Qualitative visual trichotomous assessment improves the value of fluorine-18 fluorodeoxyglucose positron emission tomography/computed tomography in predicting the prognosis of diffuse large B-cell lymphoma.

INTRODUCTION: Fluorine-18 fluorodeoxyglucose (18 F-FDG) positron emission tomography/computed tomography (PET/CT) is a powerful tool for monitoring the response of diffuse large B-cell lymphoma (DLBCL) to therapy, but the criteria to interpret PET/CT results remain under debate. We investigated the value of post-treatment PET/CT in predicting the prognosis of DLBCL patients when interpreted according to qualitative visual trichotomous assessment (QVTA) criteria compared with the Deauville criteria. METHODS: In this retrospective study, final PET/CT scans of DLBCL patients treated with rituximab-based regimens between October 2005 and November 2010 were interpreted using the Deauville and QVTA criteria. Survival curves were estimated using Kaplan-Meier analysis and compared using the log-rank test. RESULTS: A total of 253 patients were enrolled. The interpretation according to the Deauville criteria revealed that 181 patients had negative PET/CT scan results and 72 had positive results. The 3 year overall survival (OS) rate was significantly higher in patients with negative scan results than in those with positive results (91.6% vs. 57.5%, P<0.001). The 72 patients with positive scan results according to the Deauville criteria were divided into two groups by the interpretation according to the QVTA criteria: 29 had indeterminate results, and 43 had positive results. The 3 year OS rate was significantly higher in patients with indeterminate scan results than in those with positive results (91.2% vs. 33.5%, P<0.001) but was similar between patients with negative and indeterminate scan results (91.6% vs. 91.2%, P=0.921). CONCLUSIONS: Compared with the Deauville criteria, using the QVTA criteria for interpreting post-treatment PET/CT scans of DLBCL patients is likely to reduce the number of false positive results. The QVTA criteria are feasible for therapeutic outcome evaluation and can be used to guide risk-adapted therapy.

Detection of chemical vapor with high sensitivity by using the symmetrical metal-cladding waveguide-enhanced Goos-Hänchen shift.

We present a novel and simple optical structure, i.e., the symmetrical metal-cladding waveguide, in which a polymer layer is added into the guiding layer, for sensitive detection of chemical vapor by using the enhanced Goos-Hänchen (GH) shift (nearly a millimeter scale). Owing to the high sensitivity of the excited ultrahigh-order modes, the vapor-induced effect (swelling effect and refractive index change) in the polymer layer will lead to a dramatic variation of the GH shift. The detected GH shift signal is irrelevant to the power fluctuation of the incident light. The detection limit of 9.5 ppm for toluene and 28.5 ppm for benzene has been achieved.

Photodriven Radical Perfluoroalkylation-Thiolation of Unactivated Alkenes Enabled by Electron Donor-Acceptor Complex.

A clean and direct three-component radical 1,2-difunctionalization of various alkenes with perfluoroalkyl iodides and thiosulfonates enabled by the electron donor-acceptor complex has been developed under light illumination at room temperature. The approach offers a convenient and environmentally friendly route for the simultaneous incorporation of Csp3-Rf and Csp3-S bonds, affording valuable polyfunctionalized alkane derivatives containing fluorine and sulfur in satisfactory yields. Consequently, this methodology holds significant value and practicality in the field of organic synthesis.