Evaluating a technologically enhanced rehabilitation programme for wound healing in patients with coronary heart disease.

Wound healing symptoms in patients with coronary heart disease (CHD) are frequently problematic, potentially resulting in complications. In order to tackle this concern, a state-of-the-art rehabilitation programme was created, which incorporated telehealth, virtual reality and robotics in an effort to optimize wound healing in patients with coronary heart disease. The purpose of this research was to assess the efficacy of a rehabilitation programme that utilized cutting-edge technology in enhancing the outcomes of wound recovery for individuals diagnosed with (CHD). Data from 120 CHD patients who participated in this longitudinal randomized controlled trial of a specialized rehabilitation programme intended to promote wound repair were utilized in a cross-sectional analysis. Anthropometric measurements, sociodemographical factors, exercise capacity and the progression of wound recovery were gathered as data. The research participants were primarily comprised of older males from various socioeconomic backgrounds. Age, gender, BMI, socio-educational orientation, physical activity, identified regulation and identified regulation were all significant determinants of wound healing. The significance of customized strategies in cardiac rehabilitation programmes that aim to achieve favourable wound healing outcomes is underscored by these results. In conclusion, this study emphasized the importance of incorporating unique patient attributes when designing technologically advanced rehabilitation protocols aimed at facilitating wound recovery in patients with coronary heart disease. Personalized interventions that consider these variables could potentially result in improved outcomes for wound healing among this particular group of patients.

Synthesis of nitrogenase by Paenibacillus sabinae T27 in presence of high levels of ammonia during anaerobic fermentation.

Biological nitrogen fixation is usually inhibited by fixed nitrogen. Paenibacillus sabinae T27, a Gram-positive, spore-forming diazotroph, possesses high nitrogenase activity and has 3 copies of nifH (nifH, nifH2, nifH3), a copy of nifDK, and multiple nifHDK-like genes. In this study, we found that P. sabinae T27 showed nitrogenase activities not only in low (0-3 mM) concentrations of NH4+ but also in high (30-300 mM) concentrations of NH4+, no matter whether this bacterium was grown in a flask or in a fermenter on scale cultivation. qRT-PCR and western blotting analyses supported that Fe protein and MoFe protein were synthesized under both low (0-3 mM) and high (30-300 mM) concentrations of NH4+. Liquid chromatography-mass spectrometry (LC-MS) analysis revealed that MoFe protein was encoded by nifDK and Fe protein was encoded by both nifH and nifH2. The cross-reaction suggested the purified Fe and MoFe components from P. sabinae T27 grown in both nitrogen-limited and nitrogen-excess conditions were active. This is the first time to report that diazotrophs show nitrogenase activity in presence of high (30-300 mM) concentrations of NH4+. Our study will provide a clue for studying the mechanisms of nitrogen fixation in presence of the high concentration of NH4+. KEY POINTS: • P. sabinae T27 can synthesize active nitrogenase in presence of high levels of ammonia. •Fe and MoFe proteins of nitrogenase purified in absence of ammonia are the same as those purified from the high concentration of ammonia. • Fe protein is encoded by nifH and nifH2, and MoFe protein is encoded by nifDK.

Astragaloside IV enhances taxol chemosensitivity of breast cancer via caveolin-1-targeting oxidant damage.

Accumulating evidence suggests that caveolin-1 (CAV-1) is a stress-related oncotarget and closely correlated to chemoresistance. Targeting CAV-1 might be a promising strategy to improve chemosensitivity for breast cancer treatment. Astragaloside IV (AS-IV), a bioactive compound purified from Astragalus membranaceus, has been shown to exhibit multiple bioactivities, including anticancer. However, the involved molecular targets are still ambiguous. In this study, we investigated the critical role of CAV-1 in mediating the chemosensitizing effects of AS-IV to Taxol on breast cancer. We found that AS-IV could enhance the chemosensitivity of Taxol with minimal direct cytotoxicity on breast cancer cell lines MCF-7 and MDA-MB-231, as well as the nontumor mammary epithelial cell line MCF-10A. AS-IV was further demonstrated to aggravate Taxol-induced apoptosis and G2/M checkpoint arrest. The phosphorylation of mitogen-activated protein kinase (MAPK) signaling extracellular signal-regulated kinase (ERK) and c-Jun N-terminal Kinase (JNK), except p38, was also abrogated by a synergistic interaction between AS-IV and Taxol. Moreover, AS-IV inhibited CAV-1 expression in a dose-dependent manner and reversed CAV-1 upregulation induced by Taxol administration. Mechanism study further demonstrated that AS-IV treatment triggered the eNOS/NO/ONOO- pathway via inhibiting CAV-1, which led to intense oxidant damage. CAV-1 overexpression abolished the chemosensitizing effects of AS-IV to Taxol by inhibiting oxidative stress. In vivo experiments further validated that AS-IV increased Taxol chemosensitivity on breast cancer via inhibiting CAV-1 expression, followed by activation of the eNOS/NO/ONOO- pathway. Taken together, our findings not only suggested the potential of AS-IV as a promising candidate to enhance chemosensitivity, but also highlighted the significance of CAV-1 as the target to reverse cancer drug resistance.

[Research Progress of Supercontinuum Laser Spectroscopy in Biomedical Field].

Supercontinuum laser refers to the spectral broadening of an incident laser beam due to a series of nonlinear effects when the incident beam passes through a special light guide. With the development of ultrafast lasers and the photonic crystal fiber technology, the coherent and bright supercontinuum laser, generated with ultrashort pulses propagating in photonic crystal fibers, has become a kind of ideal white light source. Since supercontinuum lasers have been put into practice, their application domain is getting wider, especially for cells and bloods analysis in the biomedical field with fluorescence spectroscopy, flow cytometry, confocal microscopy, and optical coherence tomography as powerful analysis tools. Supercontinuum laser source has become one of the mail optical components in these scientific instruments that employ advanced technologies. In this paper, international research progress on supercontinuum laser sources was introduced in detail firstly, and then the development and application of supercontinuum laser spectroscopy technology in biomedical fields, such as microscopic imaging, flow cytometry instrument, fluorescence lifetime imaging microscopy, fluorescence resonance energy transfer, optical coherence tomography, and confocal microscopy was comprehensively elaborated. The requirements, schemes and research progress of supercontinuum laser spectroscopy technologies in the noncontact identification for blood products were also discussed in the paper, including studies on light fiber supercontinuum laser sources with spectra range from 400 to 2000nm, distinguishing species characteristic of bloods with supercontinuum laser spectroscopy, analyzing and establishing mathematical models based on meta databases of big data bloods spectra, determining the bloods species based on these models, and developing portable instruments for bloods spectra identification. Finally, the prospect of applications of supercontinuum laser spectroscopy in the biomedical field is discussed.

[Elemental Analysis of Rock with Remote Laser Induced Breakdown Spectroscopy].

As a kind of spectroscopic technique, the remote laser-induced breakdown spectroscopy (Remote LIBS) can qualitatively or quantitatively measure the elemental compositions of remote targets with high-power laser and focusing optical path. In this work, a Remote LIBS system was designed and established to probe the target from 2 to 10 meters. It is characterized with Cassegrain telescope structure and automatic focusing technique. Based on this system, a method to remotely retrieve the major elemental abundance of rocks is presented. With comparative experiment, the influences on spectral signal by pulse laser energy, acquisition delay time, integration time, accumulative pulse number are analyzed to find out the optimum parameters: the wavelength of laser is 1 064 nm, pulse energy is 120 mJ, delay time is 1.5 µs, integration time is 1 ms, and each spectrum data acquired with averaging 30 times detection. 48 pieces of rock specimens and 6 kinds of standard rock samples (shale, granite, andesite, basalt, gneiss and pegmatite) are selected for the experiment. As to the atomic spectra database, 8 characteristic spectral lines of the major elements (SiⅠ390.55 nm，AlⅠ394.40 nm，AlⅠ396.15 nm，CaⅡ396.85 nm，FeⅠ404.6 nm，SiⅠ500.60nm，MgⅠ518.36nm，NaⅠ589.59 nm) were extracted for analyses. Then a PLS model is constructed to quantitatively analyze the rock elements. 48 rock specimens were selected as the training sets to serve the model. The 6 standard samples were used to test the solved model. The testing results shows that the elemental abundance of Si and Al can be predicted accurately with average relative error of only 9.4% and 9.6% respectively.

[Plasma Mass Spectrometry of Sea Mountain Basalts in the Western Pacific and Its Geological Significance].

As a kind of spectroscopic technique, the remote laser-induced breakdown spectroscopy (Remote LIBS) can qualitatively or quantitatively measure the elemental compositions of remote targets with high-power laser and focusing optical path. In this work, a Remote LIBS system was designed and established to probe the target from 2 to 10 meters. It is characterized with Cassegrain telescope structure and automatic focusing technique. Based on this system, a method to remotely retrieve the major elemental abundance of rocks is presented. With comparative experiment, the influences on spectral signal by pulse laser energy, acquisition delay time, integration time, accumulative pulse number are analyzed to find out the optimum parameters: the wavelength of laser is 1 064 nm, pulse energy is 120 mJ, delay time is 1.5 µs, integration time is 1 ms, and each spectrum data acquired with averaging 30 times detection. 48 pieces of rock specimens and 6 kinds of standard rock samples (shale, granite, andesite, basalt, gneiss and pegmatite) are selected for the experiment. As to the atomic spectra database, 8 characteristic spectral lines of the major elements (SiⅠ390.55 nm，AlⅠ394.40 nm，AlⅠ396.15 nm, Ca II 396.85 nm，FeⅠ404.6 nm，SiⅠ500.60nm，MgⅠ518.36nm，NaⅠ589.59 nm) were extracted for analyses. Then a PLS model is constructed to quantitatively analyze the rock elements. 48 rock specimens were selected as the training sets to serve the model. The 6 standard samples were used to test the solved model. The testing results shows that the elemental abundance of Si and Al can be predicted accurately with average relative error of only 9.4% and 9.6% respectively.

[Identification of Animal Whole Blood Based on Near Infrared Transmission Spectroscopy].

The inspection and classification for blood products are important but complicated in import-export ports or inspection and quarantine departments. For the inspection of whole blood products, open sampling can cause pollution and virulence factors in bloods samples may even endanger inspectors. Thus non-contact classification and identification methods for whole bloods of animals are needed. Spectroscopic techniques adopted in the flowcytometry need sampling blood cells during the detection; therefore they can not meet the demand of non-contact identification and classification for whole bloods of animals. Infrared absorption spectroscopy is a technique that can be used to analyze the molecular structure and chemical bonds of detected samples under the condition of non-contact. To find a feasible spectroscopic approach of non-contact detection for the species variation in whole blood samples, a near infrared transmitted spectra (NITS, 4 497.669 - 7 506.4 cm(-1)) experiment of whole blood samples of three common animals including chickens, dogs and cats has been conducted. During the experiment, the spectroscopic resolution is 5 cm(-1), and each spectrogram is an average of 5 measured spectral data. Experimental results show that all samples have a sharp absorption peak between 5 184 and 5 215 cm(-1), and a gentle absorption peak near 7 000 cm(-1). Besides, the NITS curves of different samples of same animals are similar, and only have slight differences in the whole transmittance. A correlation coefficient (CC) is induced to distinguish the differences of the three animals' whole bloods in NITS curves, and the computed CCs between NITS curves of different samples of the same animals, are greater than 0.99, whereas CCs between NITS curves of the whole bloods of different animals are from 0.509 48 to 0.916 13. Among which CCs between NITS curves of the whole bloods of chickens and cats are from 0.857 23 to 0.912 44, CCs between NITS curves of the whole bloods of chickens and dogs are from 0.509 48 to 0.664 82, and CCs between NITS curves of the whole bloods of cats and dogs are from 0.872 75 to 0.916 13. The cat and the dog belong to the class of mammal, and the CCs between their whole bloods NITS curves are greater than those between chickens and cats, or chickens and dogs, which are hetero-class animals. Namely, the whole bloods NITS curves of the cat and the dog have higher similarity. These results of NITS provide a feasible method of non-contact identification of animal whole bloods.

[The Progress in Remote Laser-Induced Breakdown Spectroscopy].

As a kind of spectroscopic technique, the remote laser-induced breakdown spectroscopy (Remote LIBS) can measure elemental compositions of remote targets by using high-power lasers and focusing approaches. In this paper, three remote detection approaches (open path LIBS, fiber optic LIBS and compact probe fiber optic LIBS) and their system architectures are summarized and analyzed. Conventional open path LIBS, with high requirement of specifications of lasers, optical systems, spectrographs and detectors, has always been a research focus in remote testing field. Fiber optic LIBS has the advantages of simplification of optical focusing system and high collection efficiency of the plasma light. This paper reviews the progress in new techniques of LIBS, for instance Filament-LIBS techniques and LIBS combines with other spectral detection techniques, and emphatically analyzes their characteristics and advantages. These new techniques have greatly broadened the detection range of LIBS, enhanced material recognition ability of LIBS, and made a great contribution to expanding applications of remote LIBS. Latest development of applications of remote LIBS in fields of deep space exploration, hazardous material detection, pollution testing, metallurgical industries and heritage restoration is introduced in detail. With the development of laser techniques, spectral detection and calibration techniques, the detection range of remote LIBS has been expended, their application fields has been extended, and the detection precision and accuracy have been improved.

Caveolin-1 mediates chemoresistance in breast cancer stem cells via ß-catenin/ABCG2 signaling pathway.

Accumulating evidence has suggested that cancer stem cells (CSCs) are at the root of drug resistance, and recent studies have indicated that caveolin-1, a membrane transporter protein, is involved in the regulation of cancer chemoresistance and stem cell signaling. However, the current understanding of the role of caveolin-1 in breast cancer development remains controversial. Herein, we demonstrate that caveolin-1 expression was upregulated after breast cancer chemotherapy in vitro and in vivo, accompanied by co-overexpression of ß-catenin and ATP-binding cassette subfamily G member 2 (ABCG2) signaling. Additionally, breast CSCs were enriched for caveolin-1 expression. Caveolin-1 silencing sensitized breast CSCs by limiting their self-renewal ability but promoting the differentiation process. ß-catenin silencing prevented the enhanced chemoresistance of CSCs induced by caveolin-1 overexpression, indicating that ß-catenin is an essential molecule responsible for caveolin-1-mediated action. Further mechanistic investigation revealed that caveolin-1 silencing could downregulate the ß-catenin/ABCG2 pathway through glycogen synthase kinase 3 beta activation and Akt inhibition, resulting in increased ß-catenin phosphorylation and proteasomal degradation. Clinical investigation also revealed a close correlation between caveolin-1 and ß-catenin/ABCG2 signaling in breast cancer samples. Notably, caveolin-1 was highly elevated in triple-negative breast cancer, and caveolin-1 silencing significantly impaired the tumorigenicity and chemoresistance of breast CSCs in in vivo models. Overall, our study not only highlights the role of caveolin-1 in mediating the chemoresistance of breast CSCs via ß-catenin/ABCG2 regulation but also provides novel approaches for future therapies targeting CSCs.

2D Ni-Co bimetallic oxide nanosheets activate persulfate for targeted conversion of bisphenol A in wastewater into polymers.

The selective removal of targeted pollutants from complex wastewater is challenging. Herein, a novel persulfate (PS)-based advanced oxidation system equipped with a series of two-dimensional (2D) bimetallic oxide nanosheets (NSs) catalysts is developed to selectively degrade bisphenol A (BPA) within mixed pollutants via initiating nonradical-induced polymerization. Results indicate that the Ni0.60Co0.40Ox NSs demonstrate the highest catalytic efficiency among all Ni-Co NSs catalysts. Specifically, BPA degradation rate is 47.34, 27.26, and 9.72 times higher than that of 4-chlorophenol, phenol, and 2,4-dichlorophenol in the mixed solution, respectively. The lower oxidative potential of BPA in relation to the other pollutants renders it the primary target for oxidation within the PDS activation system. PDS molecules combine on the surface of Ni0.60Co0.40Ox NSs to form the surface-activated complex, triggering the generation of BPA monomer radicals through H-abstraction or electron transfer. These radicals subsequently polymerize on the surface of the catalyst through coupling reactions. Importantly, this polymerization process can occur under typical aquatic environmental conditions and demonstrates resistance to background matrices like Cl- and humic acid due to its inherent nonradical attributes. This study offers valuable insights into the targeted conversion of organic pollutants in wastewater into value-added polymers, contributing to carbon recycle and circular economy.

An integrated biological-electrocatalytic process for highly-efficient treatment of coking wastewater.

Coking wastewater is typically refractory, mainly due to its biological toxicity and complex composition. In this study, a novel integrated biological-electrocatalytic process consisting of two three-dimensional electrochemical reactors (3DERs), two biological aerated filters (BAFs), and a three-dimensional biofilm electrode reactor (3DBER) is developed for the advanced treatment of coking wastewater. 73.21% of chemical oxygen demand (COD), 38.02% of ammonium nitrogen (NH4+-N) and 91.46% of nitrate nitrogen (NO3--N) are removed by 3DERs. BAFs mainly convert NH4+-N to NO3--N through microbial nitrification. The 3DBER removes the residual NO3--N by bio-electrochemical denitrification. The integrated system can eliminate 74.72-83.27% of COD, 99.38-99.74% of NH4+-N, and 69.64-99.83% of total nitrogen from coking wastewater during the continuous operation, as well as significantly reducing the toxicity of the wastewater. The superiorities of the integrated 3DERs/BAFs/3DBER system recommend the application of such biological-electrocatalytic technology in the treatment of highly toxic wastewater.

Inhibitory effects of everolimus in combination with paclitaxel on adriamycin-resistant breast cancer cell line MDA-MB-231.

OBJECTIVE: We aimed to evaluate the therapeutic effects of paclitaxel in combination with mTOR inhibitor everolimus on adriamycin-resistant breast cancer cell line MDA-MB-231 (MDA-MB-231/ADR). MATERIALS AND METHODS: MDA-MB-231/ADR cells were treated with different concentrations of paclitaxel and everolimus. The IC50 values after 48 h of treatment were measured by the MTT assay. The apoptosis rate and cell cycle were detected by flow cytometry. The protein expressions of Akt, PI3K, mTOR, p-pI3K, p-AKT and p-mTOR were detected by Western blot. RESULTS: When paclitaxel at ≥1.56 µg/ml was used, the growth of MDA-MB-231/ADR cells was inhibited more significantly than that of control group (P < 0.05). After treatment with ≥6.25 µg/ml everolimus, the cell growth was also suppressed more significantly (P < 0.05). The IC50 values of everolimus and paclitaxel were 32.50 µg/ml and 7.80 µg/ml, respectively. The inhibition rate of paclitaxel plus everolimus was significantly enhanced with increasing paclitaxel concentration (P < 0.001). After treatment with 7.80 µg/ml paclitaxel, the two drugs had best synergistic inhibitory effects on proliferation. Compared with drugs alone, the combination significantly promoted apoptosis (P < 0.001). The paclitaxel + everolimus group had significantly more cells in the G0-G1 phase than those of control and individual drug groups (P < 0.001). Everolimus significantly decreased mTOR and p-mTOR expressions compared with those of control group (P < 0.001). Compared with everolimus alone, the combination reduced the expressions more significantly (P < 0.05). Paclitaxel decreased the expression levels of PI3K, p-PI3K and p-AKT. Compared with paclitaxel alone, the combination significantly promoted the reduction of PI3K, p-PI3K and p-AKT expressions (P < 0.05). CONCLUSION: Everolimus can enhance the effect of paclitaxel on MDA-MB-231/ADR cells, inhibit cell proliferation, induce apoptosis and arrest cell cycle in the G1 phase mainly by down-regulating the expressions of key proteins in the mTOR signaling pathway.

Caveolin-1 inhibits breast cancer stem cells via c-Myc-mediated metabolic reprogramming.

Breast cancer stem cells (BCSCs) are considered to be the root of breast cancer occurrence and progression. However, the characteristics and regulatory mechanisms of BCSCs metabolism have been poorly revealed, which hinders the development of metabolism-targeted treatment strategies for BCSCs elimination. Herein, we demonstrated that the downregulation of Caveolin-1 (Cav-1) usually occurred in BCSCs and was associated with a metabolic switch from mitochondrial respiration to aerobic glycolysis. Meanwhile, Cav-1 could inhibit the self-renewal capacity and aerobic glycolysis activity of BCSCs. Furthermore, Cav-1 loss was associated with accelerated mammary-ductal hyperplasia and mammary-tumor formation in transgenic mice, which was accompanied by enrichment and enhanced aerobic glycolysis activity of BCSCs. Mechanistically, Cav-1 could promote Von Hippel-Lindau (VHL)-mediated ubiquitination and degradation of c-Myc in BCSCs through the proteasome pathway. Notably, epithelial Cav-1 expression significantly correlated with a better overall survival and delayed onset age of breast cancer patients. Together, our work uncovers the characteristics and regulatory mechanisms of BCSCs metabolism and highlights Cav-1-targeted treatments as a promising strategy for BCSCs elimination.

Adjuvant Capecitabine With Docetaxel and Cyclophosphamide Plus Epirubicin for Triple-Negative Breast Cancer (CBCSG010): An Open-Label, Randomized, Multicenter, Phase III Trial.

PURPOSE: Standard adjuvant chemotherapy for triple-negative breast cancer (TNBC) includes a taxane and an anthracycline. Concomitant capecitabine may be beneficial, but robust data to support this are lacking. The efficacy and safety of the addition of capecitabine into the TNBC adjuvant treatment regimen was evaluated. PATIENTS AND METHODS: This randomized, open-label, phase III trial was conducted in China. Eligible female patients with early TNBC after definitive surgery were randomly assigned (1:1) to either capecitabine (3 cycles of capecitabine and docetaxel followed by 3 cycles of capecitabine, epirubicin, and cyclophosphamide) or control treatment (3 cycles of docetaxel followed by 3 cycles of fluorouracil, epirubicin, and cyclophosphamide). Randomization was centralized without stratification. The primary end point was disease-free survival (DFS). RESULTS: Between June 2012 and December 2013, 636 patients with TNBC were screened, and 585 were randomly assigned to treatment (control, 288; capecitabine, 297). Median follow-up was 67 months. The 5-year DFS rate was higher for capecitabine than for control treatment (86.3% v 80.4%; hazard ratio, 0.66; 95% CI, 0.44 to 0.99; P = .044). Five-year overall survival rates were numerically higher but not significantly improved (capecitabine, 93.3%; control, 90.7%). Overall, 39.1% of patients had capecitabine dose reductions, and 8.4% reported grade ≥ 3 hand-foot syndrome. The most common grade ≥ 3 hematologic toxicities were neutropenia (capecitabine, 136 [45.8%]; control, 118 [41.0%]) and febrile neutropenia (capecitabine, 50 [16.8%]; control, 46 [16.0%]). Safety data were similar to the known capecitabine safety profile and generally comparable between arms. CONCLUSION: Capecitabine when added to 3 cycles of docetaxel followed by 3 cycles of a 3-drug anthracycline combination containing capecitabine instead of fluorouracil significantly improved DFS in TNBC without new safety concerns.

[Effect of Astragalus mongholicus injection on proliferation and apoptosis of hormone sensitive (MCF-7) breast cancer cell lines with physiological dose E2].

OBJECTIVE: To investigate the effect of Astragalus mongholicus injection on proliferation and apoptosis in hormone sensitive breast cancer cell lines (MCF-7) with physiological dose E2. METHODS: Cell control group, TAM control group and five different dose Astragalus mongholicus injection groups with physiological dose E2 were set in the study. Proliferation of MCF-7 cells were evaluated with MTT assay, cell apoptotic rate were measured with flow cytometry, DNA ladder and cell cycle were observed. RESULTS: In physiological dose E2, Astragalus mongholicus injection inhibited MCF-7 cells proliferation at all concentration groups. As time lasting, Astragalus mongholicus injection showed better inhibitory effect than TAM (P<0.05). Among 2 x 10(-1) g/mL-2 x 10(-4) g/mL concentration, Astragalus mongholicus injection significantly increased the proliferative percent of G0/G1 and S-phase cell, decreased percent of G2-M phase cell (P<0.05) at 24 hours. After cocultured 72 hours, Astragalus mongholicus injection increased the rate of apoptosis to 16.7% at 2 x 10(-1) g/mL concentration. CONCLUSION: In some dose limit, Astragalus mongholicus injection may inhibit proliferation, induce apoptosis and interrupt caryocinesia at G0-G1 phase or S phase in hormone sensitive (MCF-7) breast cancer cell lines with physiological dose E2.

α-Lys424 Participates in Insertion of FeMoco to MoFe Protein and Maintains Nitrogenase Activity in Klebsiella oxytoca M5al.

Our previous investigation of substrates reduction catalyzed by nitrogenase suggested that α-Ile423 of MoFe protein possibly functions as an electron transfer gate to Mo site of active center-"FeMoco". Amino acid residue α-Lys424 connects directly to α-Ile423, and they are located in the same α-helix (α423-431). In the present study, function of α-Lys424 was investigated by replacing it with Arg (alkaline, like Lys), Gln (neutral), Glu (acidic), and Ala (neutral) through site-directed mutagenesis and homologous recombination. The mutants were, respectively, termed 424R, 424Q, 424E, and 424A. Studies of diazotrophic cell growth, cytological, and enzymatic properties indicated that none of the substitutions altered the secondary structure of MoFe protein, or normal expression of nifA, nifL, and nifD. Substitution of alkaline amino acid (i.e., 424R) maintained acetylene (C2H2) and proton (H+) reduction activities at normal levels similar to that of wild-type (WT), because its FeMoco content did not reduce. In contrast, substitution of acidic or neutral amino acid (i.e., 424Q, 424E, 424A) impaired the catalytic activity of nitrogenase to varying degrees. Combination of MoFe protein structural simulation and the results of a series of experiments, the function of α-Lys424 in ensuring insertion of FeMoco to MoFe protein was further confirmed, and the contribution of α-Lys424 in maintaining low potential of the microenvironment causing efficient catalytic activity of nitrogenase was demonstrated.

Combined Assembly and Targeted Integration of Multigene for Nitrogenase Biosynthetic Pathway in Saccharomyces cerevisiae.

Biological nitrogen fixation, a process unique to diazotrophic prokaryote, is catalyzed by the nitrogenase complex. There has been a long-standing interest in reconstituting a nitrogenase biosynthetic pathway in a eukaryotic host with the final aim of developing N2-fixing cereal crops. In this study, we report that a nitrogenase biosynthetic pathway (∼38 kb containing 15 genes) was assembled in two individual one-step methods via in vivo assembly and integrated at δ and HO sites in Saccharomyces cerevisiae chromosome. Of the 15 genes, 11 genes (nifB, nifH, nifD, nifK, nifE, nifN, nifX, hesA, nifV, groES, groEL) were from Paenibacillus polymyxa WLY78 and 4 genes (nifS, nifU, nifF, nifJ) were from Klebsiella oxytoca. The 15-gene nitrogenase biosynthetic pathway was correctly assembled and transcribed in the recombinant S. cerevisiae. The NifDK tetramer with an identical molecular weight as that of P. polymyxa was formed in yeast and the expressed NifH exhibited the activity of Fe protein. This study demonstrates that it will be possible to produce active nitrogenase in eukaryotic hosts.

[Effect of Zirconium-Modified Zeolite Addition on Migration and Transformation of Phosphorus in River Sediments Under Static and Hydrodynamic Disturbance Conditions].

In this study, the effect of the addition of zirconium-modified zeolite (ZrMZ) on the migration and transformation of phosphorus (P) in river sediments under static and hydrodynamic disturbance conditions was studied using sediment core incubation experiments. Results showed that, whether under static or hydrodynamic disturbance condition, the ZrMZ amendment suppressed the release of SRP from sediments into the overlying water. Furthermore, the addition of ZrMZ to the upper sediment (0-10 mm) not only resulted in the decrease of the dissoluble reactive P (SRP) concentration in the overlying water at a depth of 0-30 mm, but also led to the decrease of the diffusion flux of SRP from the pore water to the overlying water across the sediment-water interface (SWI). In addition, the ZrMZ amendment induced the transformation of the redox-sensitive P (BD-P) and HCl extractable P (HCl-P) into the metal oxide-bound P (NaOH-rP) and residual P (Res-P), thus resulting in the reduction of mobile P (sum of NH4Cl extractable P and BD-P) in the top 10 mm of sediment. In addition, the addition of ZrMZ into the top 10 mm of sediment resulted in reduction of the content of mobile P in 10-20 mm of sediment. Furthermore, the effect of ZrMZ addition on the migration and transformation of P in sediments under hydrodynamic disturbance condition had a certain difference from that under static condition. The presence of hydrodynamic disturbance enhanced the immobilization efficiency of SRP in the pore water at a depth of 0-20 mm by the ZrMZ amendment, and also increased the reduction efficiency of the SRP diffusion flux from the pore water to the overlying water across the SWI by the ZrMZ amendment. However, the efficiency of the control of SRP release from sediments to the overlying water by the ZrMZ amendment was slightly reduced by the hydrodynamic disturbance. The reductions of mobile P in the top sediment, SRP in the pore water as well as the diffusion flux of SRP from the pore water to the overlying water across the SWI played a key role in the control of SRP release from sediments to the overlying water by the ZrMZ amendment. Results of this work indicate that ZrMZ is a very promising amendment for the control of SRP release from river sediments under static and hydrodynamic disturbance conditions.

Betulinic Acid Suppresses Breast Cancer Metastasis by Targeting GRP78-Mediated Glycolysis and ER Stress Apoptotic Pathway.

Targeting aberrant metabolism is a promising strategy for inhibiting cancer growth and metastasis. Research is now geared towards investigating the inhibition of glycolysis for anticancer drug development. Betulinic acid (BA) has demonstrated potent anticancer activities in multiple malignancies. However, its regulatory effects on glycolysis and the underlying molecular mechanisms are still unclear. BA inhibited invasion and migration of highly aggressive breast cancer cells. Moreover, BA could suppress aerobic glycolysis of breast cancer cells presenting as a reduction of lactate production, quiescent energy phenotype transition, and downregulation of aerobic glycolysis-related proteins. In this study, glucose-regulated protein 78 (GRP78) was also identified as the molecular target of BA in inhibiting aerobic glycolysis. BA treatment led to GRP78 overexpression, and GRP78 knockdown abrogated the inhibitory effect of BA on glycolysis. Further studies demonstrated that overexpressed GRP78 activated the endoplasmic reticulum (ER) stress sensor PERK. Subsequent phosphorylation of eIF2α led to the inhibition of ß-catenin expression, which resulted in the inhibition of c-Myc-mediated glycolysis. Coimmunoprecipitation assay revealed that BA interrupted the binding between GRP78 and PERK, thereby initiating the glycolysis inhibition cascade. Finally, the lung colonization model validated that BA inhibited breast cancer metastasis in vivo, as well as suppressed the expression of aerobic glycolysis-related proteins. In conclusion, our study not only provided a promising drug for aerobic glycolysis inhibition but also revealed that GRP78 is a novel molecular link between glycolytic metabolism and ER stress during tumor metastasis.

Broadleaf Mahonia attenuates granulomatous lobular mastitis­associated inflammation by inhibiting CCL­5 expression in macrophages.

Granulomatous lobular mastitis (GLM) is a type of chronic mammary inflammation with unclear etiology. Currently systematic corticosteroids and methitrexate are considered as the main drugs for GLM treatment, but a high toxicity and risk of recurrence greatly limit their application. It is therefore an urgent requirement that safe and efficient natural drugs are found to improve the GLM prognosis. Broadleaf Mahonia (BM) is a traditional Chinese herb that is believed to have anti­inflammatory properties according to ancient records of traditional Chinese medicine. The present study investigated this belief and demonstrated that BM significantly inhibited the expression of interleukin­1ß (IL­1ß), IL­6, cyclooxygenase­2 and inducible nitric oxide synthase in RAW264.7 cells, but had little influence on the cell viability, cell cycle and apoptosis. Meanwhile, the lipopolysaccharide­induced elevation of reactive oxygen species and nitric oxide was also blocked following BM treatment, accompanied with decreased activity of nuclear factor­κB and MAPK signaling. A cytokine array further validated that BM exhibited significant inhibitory effects on several chemoattractants, including chemokine (C­C motif) ligand (CCL)­2, CCL­3, CCL­5 and secreted tumor necrosis factor receptor 1, among which CCL­5 exhibited the highest inhibition ratio in cell and clinical GLM specimens. Collectively, the results show that BM is a novel effective anti­inflammatory herb in vitro and ex vivo, and that CCL­5 may be closely associated with GLM pathogenesis.