Phosphorylation Modification, Structural Characterization, Antioxidant and DNA Protection Capacities of Polysaccharides from Asarum Sieboldii Miq.

Polysaccharide from Asarum sieboldii Miq (ASP) was extracted and five phosphorylation polysaccharides with different degree of substitution were obtained, namely ASPP1, ASPP2, ASPP3, ASPP4, and ASPP5 (ASPPs). The physical and chemical structure and biological activities were studied. The results suggested that the carbohydrate and protein content were reduced while uronic acid was increased after phosphorylation modification. The molecular weight of ASPPs was significantly lower than that of ASP. ASPPs were acidic heteropolysaccharides mainly composed of galacturonic acid, galactose, glucose, fructose, and arabinose. The UV-vis spectrum indicated that the polysaccharides did not contain nucleic acid or protein after modification. The Fourier transform infrared spectrum demonstrated that ASPPs contained characteristic absorption peaks of P=O and P-O-C near 1270 and 980 cm-1 . ASPPs presented a triple helix conformation, but it was not presented in ASP. The scanning electron microscopy analysis showed that the surface topography and particle structure of ASP were different after modification. Compared with ASP, ASPPs enhanced the activity to scavenge DPPH and ABTS free radicals and possessed more protective ability to DNA oxidation caused by OH⋅, GS⋅, and AAPH free radicals. These results suggest that chemical modification is beneficial for the exploitation and utilization of natural polysaccharides.

Transcriptomic-Proteomic Analysis Revealed the Regulatory Mechanism of Peanut in Response to Fusarium oxysporum.

Peanut Fusarium rot, which is widely observed in the main peanut-producing areas in China, has become a significant factor that has limited the yield and quality in recent years. It is highly urgent and significant to clarify the regulatory mechanism of peanuts in response to Fusarium oxysporum. In this study, transcriptome and proteome profiling were combined to provide new insights into the molecular mechanisms of peanut stems after F. oxysporums infection. A total of 3746 differentially expressed genes (DEGs) and 305 differentially expressed proteins (DEPs) were screened. The upregulated DEGs and DEPs were primarily enriched in flavonoid biosynthesis, circadian rhythm-plant, and plant-pathogen interaction pathways. Then, qRT-PCR analysis revealed that the expression levels of phenylalanine ammonia-lyase (PAL), chalcone isomerase (CHI), and cinnamic acid-4-hydroxylase (C4H) genes increased after F. oxysporums infection. Moreover, the expressions of these genes varied in different peanut tissues. All the results revealed that many metabolic pathways in peanut were activated by improving key gene expressions and the contents of key enzymes, which play critical roles in preventing fungi infection. Importantly, this research provides the foundation of biological and chemical analysis for peanut disease resistance mechanisms.

Targeting the KRT16-vimentin axis for metastasis in lung cancer.

Lung cancer is the most diagnosed malignant cancer and the leading cause of cancer-related deaths worldwide, with advanced stage and metastasis being a major issue. The mechanism leading to metastasis is not yet understood. Here, we found that KRT16 is upregulated in metastatic lung cancer tissues and correlated with poor overall survival. Knockdown of KRT16 inhibits metastasis of lung cancer both in vitro and in vivo. Mechanistically, KRT16 interacts with vimentin, and depletion of KRT16 leads to downregulation of vimentin. KRT16 acquired its oncogenic ability by stabilizing vimentin, and vimentin is required for KRT16-driven metastasis. FBXO21 mediates the polyubiquitination and degradation of KRT16, and vimentin inhibits KRT16 ubiquitination and degradation by impairing its interaction with FBXO21. Significantly, IL-15 inhibits metastasis of lung cancer in a mouse model through upregulation of FBXO21, and the level of IL-15 in circulating serum was significantly higher in nonmetastatic lung cancer patients than in metastatic patients. Our findings indicate that targeting the FBXO21/KRT16/vimentin axis may benefit lung cancer patients with metastasis.

Gastrointestinal perforation associated with novel antineoplastic agents: A real-world study based on the FDA Adverse Event Reporting System.

Purpose: Gastrointestinal perforation (GIP) is a fatal adverse event (AE). The AE of GIP induced by novel antineoplastic agents has attracted attention recently. We aimed to explore the AE signals of GIP related to novel antineoplastic agents comprehensively based on the FDA Adverse Event Reporting System (FAERS). Methods: The FAERS database containing 71 quarters of records was used for analysis. Reporting odds ratio (ROR), information component (IC), and empirical Bayesian geometric mean (EBGM) were utilized to evaluate the signals of GIP associated with novel antineoplastic drugs. Standardization of drug names was by employing MedEx-UIMA software and Python. Data analysis and visualization were performed using MySQL Workbench and R software. Results: After cleaning and handling the data, 5226 GIP cases were identified that were associated with new antineoplastic medications, where these agents were the main suspected contributors. A total of 37 novel antineoplastic drugs were detected with signals of GIP for ROR and IC. Only 22 drugs showed statistically significant signals for EBGM. We found the GIP signals of 22 novel antineoplastic drugs overlapped for the 3 indicators, including anti-vascular endothelial growth factor/vascular endothelial growth factor receptor, anti-endothelial growth factor receptor, immune checkpoint inhibitors, and so on. Conclusion: The potential risk of GIP associated with several novel antineoplastic agents was identified through data mining, which provided valuable information on the safety risks associated with GIP among these drugs. The potential threat of GIP should be recognized and managed properly when using these novel antineoplastic agents.

Developmental toxicity of TCBPA on the nervous and cardiovascular systems of zebrafish (Danio rerio): A combination of transcriptomic and metabolomics.

Tetrachlorobisphenol A (TCBPA), a widely used halogenated flame retardant, is frequently detected in environmental compartments and human samples. However, unknown developmental toxicity and mechanisms limit the entire understanding of its effects. In this study, zebrafish (Danio rerio) embryos were exposed to various concentrations of TCBPA while a combination of transcriptomics, behavioral and biochemical analyzes as well as metabolomics were applied to decipher its toxic effects and the potential mechanisms. We found that TCBPA could interfere with nervous and cardiovascular development through focal adhesion and extracellular matrix-receptor (ECM-receptor) interaction pathways through transcriptomic analysis. Behavioral and biochemical analysis results indicated abnormal swimming behavior of zebrafish larvae. Morphological observations revealed that TCBPA could cause the loss of head blood vessels. Metabolomic analysis showed that arginine-related metabolic pathways were one of the main pathways leading to TCBPA developmental toxicity. Our study demonstrated that by using omics, TCBPA was shown to have neurological and cardiovascular developmental toxicity and the underlying mechanisms were uncovered and major pathways identified.

Bioglass promotes wound healing by inhibiting endothelial cell pyroptosis through regulation of the connexin 43/reactive oxygen species (ROS) signaling pathway.

Bioactive glass (BG) has recently shown great promise in soft tissue repair, especially in wound healing; however, the underlying mechanism remains unclear. Pyroptosis is a novel type of programmed cell death that is involved in various traumatic injury diseases. Here, we hypothesized that BG may promote wound healing through suppression of pyroptosis. To test this scenario, we investigated the possible effect of BG on pyroptosis in wound healing both in vivo and in vitro. This study showed that BG can accelerate wound closure, granulation formation, collagen deposition, and angiogenesis. Moreover, western blot analysis and immunofluorescence staining revealed that BG inhibited the expression of pyroptosis-related proteins in vivo and in vitro. In addition, while BG regulated the expression of connexin43 (Cx43), it inhibited reactive oxygen species (ROS) production. Cx43 activation and inhibition experiments further indicate that BG inhibited pyroptosis in endothelial cells by decreasing Cx43 expression and ROS levels. Taken together, these studies suggest that BG promotes wound healing by inhibiting pyroptosis via Cx43/ROS signaling pathway.

Sesquiterpenes and Monoterpenes from the Leaves and Stems of Illicium simonsii and Their Antibacterial Activity.

Two undescribed ether derivatives of sesquiterpenes, 1-ethoxycaryolane-1, 9ß-diol (1) and 2-ethoxyclovane-2ß, 9α-diol (3), and one new monoterpene glycoside, p-menthane-1α,2α,8-triol-4-O-ß-D-glucoside (5), were obtained, together with eight known compounds from the stems and leaves of I. simonsii. Their structures were elucidated by spectroscopic methods. Compounds 1-11 were evaluated for their potency against Staphylococcus aureus and clinical methicillin-resistant S. aureus (MRSA). Among them, compound 3 was weakly active against S. aureus (MIC = 128 µg/mL), and compounds 6 and 7 exhibited good antibacterial activity against S. aureus and MRSA (MICs = 2-8 µg/mL). A primary mechanism study revealed that compounds 6 and 7 could kill bacteria by destroying bacterial cell membranes. Moreover, compounds 6 and 7 were not susceptible to drug resistance development.

Tough hybrid microgel-reinforced hydrogels dependent on the size and modulus of the microgels.

Microgel-reinforced (MR) hydrogels are tough hydrogels with dispersed rigid microgels embedded in a continuous soft matrix. MR gels have the great potential to provide not only mechanical toughness but also the desired functional matrix by incorporation of various functional microgels. Understanding the toughening mechanism of the MR hydrogels is critical for the rational design of the desired functionally tough MR gels. However, our current knowledge of the toughening mechanism of MR gels mainly comes from the MR hydrogels with both chemically crosslinked dispersed microgels and a continuous matrix. Little is known about the hybrid MR gels with physically crosslinked microgels embedded in a chemically crosslinked matrix. Herein, we synthesize such hybrid MR hydrogels with the ionic crosslinked calcium alginate microgels incorporated into the chemically crosslinked polyacrylamide (PAAm) matrix. The alginate microgels show strong size and modulus effects on the toughening enhancement: the larger microgels could toughen the MR gels more than the small ones, and the microgels with medium modulus could maximize the toughness of the MR gels. By comparison of the mechanical performances of the MR and the corresponding double network (DN) hydrogels, we have proposed that the hybrid MR gels may have the same toughening mechanism as the bulk DN gel. This work tries to better understand the structure-property relationships of both MR and DN gels and help in the design of more functionally tough MR gels with the desired properties.

Targetability of cervical cancer by magnetic resonance-guided high-intensity focused ultrasound (MRgHIFU)-mediated hyperthermia (HT) for patients receiving radiation therapy.

PURPOSE: To evaluate the targetability of late-stage cervical cancer by magnetic resonance-guided high-intensity focused ultrasound (MRgHIFU)-induced hyperthermia (HT) as an adjuvant to radiation therapy (RT). METHODS: Seventy-nine cervical cancer patients (stage IIIB-IVA) who received RT with lesions visible on positron emission tomography-computed tomography (PET-CT) were retrospectively analyzed for targetability using a commercially-available HT-capable MRgHIFU system. Targetability was assessed for both primary targets and/or any metastatic lymph nodes using both posterior (supine) and anterior (prone) patient setups relative to the transducer. Thirty-four different angles of rotation along subjects' longitudinal axis were analyzed. Targetability was categorized as: (1) Targetable with/without minimal intervention; (2) Not targetable. To determine if any factors could be used for prospective screening of patients, potential associations between demographic/anatomical factors and targetability were analyzed. RESULTS: 72.15% primary tumors and 33.96% metastatic lymph nodes were targetable from at least one angle. 49.37% and 39.24% of primary tumors could be targeted with patient laying in supine and prone positions, respectively. 25°-30° rotation and 0° rotation had the highest rate of the posterior and anterior targetability, respectively. The ventral depth of the tumor and its distance to the coccyx were statistically correlated with the anterior and posterior targetability, respectively. CONCLUSION: Most late-stage cervical cancer primaries were targetable by MRgHIFU HT requiring either no/minimal intervention. A rotation of 0° or 25°-30° relative to the transducer might benefit anterior and posterior targetability, respectively. Certain demographic/anatomic parameters might be useful in screening patients for treatability.

Liquid Metal-Based Soft Microfluidics.

Motivated by the increasing demand of wearable and soft electronics, liquid metal (LM)-based microfluidics has been subjected to tremendous development in the past decade, especially in electronics, robotics, and related fields, due to the unique advantages of LMs that combines the conductivity and deformability all-in-one. LMs can be integrated as the core component into microfluidic systems in the form of either droplets/marbles or composites embedded by polymer materials with isotropic and anisotropic distribution. The LM microfluidic systems are found to have broad applications in deformable antennas, soft diodes, biomedical sensing chips, transient circuits, mechanically adaptive materials, etc. Herein, the recent progress in the development of LM-based microfluidics and their potential applications are summarized. The current challenges toward industrial applications and future research orientation of this field are also summarized and discussed.

Correction to: LncRNA MEG3 promotes melanoma growth, metastasis and formation through modulating miR-21/E-cadherin axis.

[This corrects the article DOI: 10.1186/s12935-019-1087-4.].

LncRNA MEG3 promotes melanoma growth, metastasis and formation through modulating miR-21/E-cadherin axis.

BACKGROUND: Melanoma is the most aggressive type of skin cancer with high mortality rate and poor prognosis. lncRNA MEG3, a tumor suppressor, is closely related to the development of various cancers. However, the role of lncRNA MEG3 in melanoma has seldom been studied. METHODS: RT-PCR was used to examine the expressions of lncRNA MEG3 and E-cadherin in melanoma patients and cell lines. Then, the biological functions of lncRNA MEG3 and E-cadherin were demonstrated by transfecting lncRNA MEG3-siRNA, lncRNA MEG3-overexpression, E-cadherin-siRNA and E-cadherin-overexpression plasmids in melanoma cell lines. Moreover, CCK8 assay and colony formation assay were utilized to assess the cell proliferation; Transwell assay was performed to evaluate the cell invasive ability; and tumor xenografts in nude mice were applied to test the tumor generation. Additionally, the target interactions among lncRNA MEG3, miR-21 and E-cadherin were determined by dual luciferase reporter assay. Finally, RT-PCR and WB were further conducted to verify the regulatory roles among lncRNA MEG3, miR-21 and E-cadherin. RESULTS: The clinical data showed that lncRNA MEG3 and E-cadherin expressions were both declined in carcinoma tissues as compared with their para-carcinoma tissues. Moreover, lncRNA MEG3 and E-cadherin expressions in B16 cells were also higher than those in A375 and A2058 cells. Subsequently, based on the differently expressed lncRNA MEG3 and E-cadherin in these human melanoma cell lines, we chose B16, A375 and A2058 cells for the following experiments. The results demonstrated that lncRNA MEG3 could suppress the tumor growth, tumor metastasis and formation; and meanwhile E-cadherin had the same effects on tumor growth, tumor metastasis and formation. Furthermore, the analysis of Kaplan-Meier curves also confirmed that there was a positive correlation between lncRNA MEG3 and E-cadherin. Ultimately, dual luciferase assays were further used to verify that lncRNA MEG3 could directly target miR-21 which could directly target E-cadherin in turn. Additionally, the data of RT-PCR and WB revealed that knockdown of lncRNA MEG3 in B16 cells inhibited miR-21 expression and promoted E-cadherin expression, but overexpression of lncRNA MEG3 in A375 and A2058 cells presented completely opposite results. CONCLUSION: Our findings indicated that lncRNA MEG3 might inhibit the tumor growth, tumor metastasis and formation of melanoma by modulating miR-21/E-cadherin axis.

Expression regulation of multiple key genes to improve L-threonine in Escherichia coli.

BACKGROUND: Escherichia coli is an important strain for L-threonine production. Genetic switch is a ubiquitous regulatory tool for gene expression in prokaryotic cells. To sense and regulate intracellular or extracellular chemicals, bacteria evolve a variety of transcription factors. The key enzymes required for L-threonine biosynthesis in E. coli are encoded by the thr operon. The thr operon could coordinate expression of these genes when L-threonine is in short supply in the cell. RESULTS: The thrL leader regulatory elements were applied to regulate the expression of genes iclR, arcA, cpxR, gadE, fadR and pykF, while the threonine-activating promoters PcysH, PcysJ and PcysD were applied to regulate the expression of gene aspC, resulting in the increase of L-threonine production in an L-threonine producing E. coli strain TWF001. Firstly, different parts of the regulator thrL were inserted in the iclR regulator region in TWF001, and the best resulting strain TWF063 produced 16.34 g L-threonine from 40 g glucose after 30 h cultivation. Secondly, the gene aspC following different threonine-activating promoters was inserted into the chromosome of TWF063, and the best resulting strain TWF066 produced 17.56 g L-threonine from 40 g glucose after 30 h cultivation. Thirdly, the effect of expression regulation of arcA, cpxR, gadE, pykF and fadR was individually investigated on L-threonine production in TWF001. Finally, using TWF066 as the starting strain, the expression of genes arcA, cpxR, gadE, pykF and fadR was regulated individually or in combination to obtain the best strain for L-threonine production. The resulting strain TWF083, in which the expression of seven genes (iclR, aspC, arcA, cpxR, gadE, pykF, fadR and aspC) was regulated, produced 18.76 g L-threonine from 30 g glucose, 26.50 g L-threonine from 40 g glucose, or 26.93 g L-threonine from 50 g glucose after 30 h cultivation. In 48 h fed-batch fermentation, TWF083 could produce 116.62 g/L L-threonine with a yield of 0.486 g/g glucose and productivity of 2.43 g/L/h. CONCLUSION: The genetic engineering through the expression regulation of key genes is a better strategy than simple deletion of these genes to improve L-threonine production in E. coli. This strategy has little effect on the intracellular metabolism in the early stage of the growth but could increase L-threonine biosynthesis in the late stage.

Characterization of magnetic resonance-guided high-intensity focused ultrasound (MRgHIFU)-induced large-volume hyperthermia in deep and superficial targets in a porcine model.

PURPOSE: To characterize temperature fields and tissue damage profiles of large-volume hyperthermia (HT) induced by magnetic resonance-guided high-intensity focused ultrasound (MRgHIFU) in deep and superficial targets in vivo in a porcine model. METHODS: Nineteen HT sessions were performed in vivo with a commercial MRgHIFU system (Sonalleve® V2, Profound Medical Inc., Mississauga, ON, Canada) in hind leg muscles of eight pigs with temperature fields of cross-sectional diameter of 58-mm. Temperature statistics evaluated in the target region-of-interest (tROI) included accuracy, temporal variation, and uniformity. The impact of the number and location of imaging planes for feedback-based temperature control were investigated. Temperature fields were characterized by time-in-range (TIR, the duration each voxel stays within 40-45 °C) maps. Tissue damage was characterized by contrast-enhanced MRI, and macroscopic and histopathological analysis. The performance of the Sonalleve® system was benchmarked against a commercial phantom. RESULTS: Across all HT sessions, the mean difference between the average temperature (Tavg) and the desired temperature was -0.4 ± 0.5 °C; the standard deviation of temperature 1.2 ± 0.2 °C; the temporal variation of Tavg for 30-min HT was 0.6 ± 0.2 °C, and the temperature uniformity was 1.5 ± 0.2 °C. A difference of 2.2-cm (in pig) and 1.5-cm (in phantom) in TIR dimensions was observed when applying feedback-based plane(s) at different locations. Histopathology showed 62.5% of examined HT sessions presenting myofiber degeneration/necrosis within the target volume. CONCLUSION: Large-volume MRgHIFU-mediated HT was successfully implemented and characterized in a porcine model in deep and superficial targets in vivo with heating distributions modifiable by user-definable parameters.

The Brassicaceae-specific secreted peptides, STMPs, function in plant growth and pathogen defense.

Low molecular weight secreted peptides have recently been shown to affect multiple aspects of plant growth, development, and defense responses. Here, we performed stepwise BLAST filtering to identify unannotated peptides from the Arabidopsis thaliana protein database and uncovered a novel secreted peptide family, secreted transmembrane peptides (STMPs). These low molecular weight peptides, which consist of an N-terminal signal peptide and a transmembrane domain, were primarily localized to extracellular compartments but were also detected in the endomembrane system of the secretory pathway, including the endoplasmic reticulum and Golgi. Comprehensive bioinformatics analysis identified 10 STMP family members that are specific to the Brassicaceae family. Brassicaceae plants showed dramatically inhibited root growth upon exposure to chemically synthesized STMP1 and STMP2. Arabidopsis overexpressing STMP1, 2, 4, 6, or 10 exhibited severely arrested growth, suggesting that STMPs are involved in regulating plant growth and development. In addition, in vitro bioassays demonstrated that STMP1, STMP2, and STMP10 have antibacterial effects against Pseudomonas syringae pv. tomato DC3000, Ralstonia solanacearum, Bacillus subtilis, and Agrobacterium tumefaciens, demonstrating that STMPs are antimicrobial peptides. These findings suggest that STMP family members play important roles in various developmental events and pathogen defense responses in Brassicaceae plants.

Electric Actuation of Liquid Metal Droplets in Acidified Aqueous Electrolyte.

The electric actuation of room-temperature liquid metals, such as Galinstan (gallium-indium-tin), has largely been conducted in alkaline electrolyte. Addition of surface-active anions and a proper acidic pH are expected to influence the interfacial tension of the liquid metal due to a high surface charge density. Hence, it should be possible to actuate liquid metals in such acidic environments. To ascertain this, at first, the dependence of the interfacial tension of Galinstan in NaOH, acidified KI, and acidified NaCl electrolyte on the concentration of the surface-active anions OH-, I-, and Cl-, respectively, were studied. Subsequently, a systematic study of the actuation of Galinstan in acidified KI electrolyte was executed and compared to actuation in alkaline medium. In the presence of HCl and acidified NaCl electrolyte, the interfacial tension of Galinstan is only marginally altered, while acidified KI solution reduced the interfacial tension of Galinstan significantly from 470.8 ± 1.4 (no KI) to 370.6 ± 4.1 mN/m (5 M KI) due to the high surface charge density of the electric double layer. Therefore, in acidified electrolyte in the presence of surface-active anions, the electrically actuated motion of LM can be realized. In particular, the actuation of Galinstan achieves a higher average and maximum speed at lower applied voltage and power consumption for acidified KI electrolyte. The formation of high surface charge density in acidified environments signifies a paradigm shift and opens up new possibilities to tune interfacial tension and controlled LM droplet motion of room-temperature liquid metals.

Liquid Metal-Mediated Mechanochemical Polymerization.

Mechanically controlled polymerization that employs the mechanical energy to fabricate novel synthetic materials has attracted considerable interest. However, only a few examples have been achieved so far, owing to the limited choices of materials and strategies. Herein, a versatile, liquid metal (LM)-mediated mechanochemical polymerization method (LMMMP) is developed for the air-compatible, robust preparation of polymers in an aqueous solution. This method involves the simultaneous disruption of bulk LMs into micro- and nanodroplets and the combination of monomers into polymers during ultrasonic irradiation. The pristine and reactive LM surface continuously generated by ultrasound endows this polymerization method with excellent oxygen tolerance, high reaction rate, and the ability to produce polymers with high molecular weight from a wide variety of water-soluble monomers. Besides, LM droplets are readily reclaimed and reused for polymerization. The authors envision that the LMMMP promotes the utilization of mechanical energy for the synthesis of functional polymers, constitutes a novel fabrication approach for polymer-LM nanocomposites, and provides new insight into the design of LM-based platforms for polymerization.

Feasibility and safety assessment of magnetic resonance-guided high-intensity focused ultrasound (MRgHIFU)-mediated mild hyperthermia in pelvic targets evaluated using an in vivo porcine model.

Purpose: To evaluate the feasibility and assess safety parameters of magnetic resonance-guided high-intensity focused ultrasound (MRgHIFU)-mediated hyperthermia (HT; heating to 40-45 °C) in various pelvic targets in a porcine model in vivo.Methods: Thirteen HT treatments were performed in six pigs with a commercial MRgHIFU system (Sonalleve V2, Profound Medical Inc., Mississauga, Canada) to muscle adjacent to the ventral/dorsal bladder wall and uterus to administer 42 °C (±1°) for 30 min (±5%) using an 18-mm target diameter and 100 W power. Feasibility was assessed using accuracy, uniformity, and MR-thermometry performance-based metrics. Safety parameters were assessed for tissues in the targets and beam-path by contrast-enhanced MRI, gross-pathology and histopathology.Results: Across all HT sessions, the mean difference between average temperature (Tavg) and the target temperature within the target region-of-interest (tROI, the cross-section of the heated volume at focal depth) was 0.51 ± 0.33 °C. Within the tROI, the temperature standard deviation averaged 1.55 ± 0.31 °C, the average 30-min Tavg variation was 0.80 ± 0.17 °C, and the maximum difference between Tavg and the 10th- or 90th-percentile temperature averaged 2.01 ± 0.44 °C. The average time to reach ≥41 °C and cool to ≤40 °C within the tROI at the beginning and end of treatment was 47.25 ± 27.47 s and 66.37 ± 62.68 s, respectively. Compared to unheated controls, no abnormally-perfused tissue or permanent damage was evident in the MR images, gross pathology or histological analysis.Conclusions: MRgHIFU-mediated HT is feasible and safety assessment is satisfactory for treating an array of clinically-mimicking pelvic geometries in a porcine model in vivo, implying the technique may have utility in treating pelvic targets in human patients.

Developing an l-threonine-producing strain from wild-type Escherichia coli by modifying the glucose uptake, glyoxylate shunt, and l-threonine biosynthetic pathway.

Wild-type Escherichia coli MG1655 usually does not accumulate l-threonine. In this study, the effects of 13 genes related to the glucose uptake, glycolysis, TCA cycle, l-threonine biosynthesis, or their regulation on l-threonine accumulation in E. coli MG1655 were investigated. Sixteen E. coli mutant strains were constructed by chromosomal deletion or overexpression of one or more genes of rsd, ptsG, ptsH, ptsI, crr, galP, glk, iclR, and gltA; the plasmid pFW01-thrA*BC-rhtC harboring the key genes for l-threonine biosynthesis and secretion was introduced into these mutants. The analyses on cell growth, glucose consumption, and l-threonine production of these recombinant strains showed that most of these strains could accumulate l-threonine, and the highest yield was obtained in WMZ016/pFW01-thrA*BC-rhtC. WMZ016 was derived from MG1655 by deleting crr and iclR and enhancing the expression of gltA. WMZ016/pFW01-thrA*BC-rhtC could produce 17.98 g/L l-threonine with a yield of 0.346 g/g glucose, whereas the control strain MG1655/pFW01-thrA*BC-rhtC could only produce 0.68 g/L l-threonine. In addition, WMZ016/pFW01-thrA*BC-rhtC could tolerate the high concentration of glucose and produced no detectable by-products; therefore, it should be an ideal platform strain for further development. The results indicate that manipulating the glucose uptake and TCA cycle could efficiently increase l-threonine production in E. coli.

Deletion of arcA, iclR, and tdcC in Escherichia coli to improve l-threonine production.

l-Threonine is an important amino acid supplemented in food, medicine, or feed. Starting from glucose, l-threonine production in Escherichia coli involves the glycolysis, TCA cycle, and the l-threonine biosynthetic pathway. In this study, how the l-threonine production in an l-threonine producing E. coli TWF001 is controlled by the three regulators ArcA, Cra, and IclR, which control the expression of genes involved in the glycolysis and TCA cycle, has been investigated. Ten mutant strains were constructed from TWF001 by different combinations of deletion or overexpression of arcA, cra, iclR, and tdcC. l-Threonine production was increased in the mutants TWF015 (ΔarcAΔcra), TWF016 (ΔarcAPcra::Ptrc), TWF017 (ΔarcAΔiclR), TWF018 (ΔarcAΔiclRΔtdcC), and TWF019 (ΔarcAΔcraΔiclRΔtdcC). Among these mutant strains, the highest l-threonine production (26.0 g/L) was obtained in TWF018, which was a 109.7% increase compared with the control TWF001. In addition, TWF018 could consume glucose more efficiently than TWF001 and produce less acetate. The results suggest that deletion of arcA, iclR, and tdcC could efficiently increase l-threonine production in E. coli.