Clear Cell Renal Cell Carcinoma: Characterizing the Phenotype of Von Hippel-Lindau Mutation Using MRI.

BACKGROUND: The von Hippel-Lindau (VHL) mutation is an important alteration in clear cell renal cell carcinoma (ccRCC); however, its imaging phenotype remains unclear. PURPOSE: To investigate whether MRI features can reflect the VHL mutation status. STUDY TYPE: Retrospective. FIELD STRENGTH/SEQUENCE: 3 T/fast spin echo T2-weighted, spin-echo echo planar diffusion-weighted, gradient recalled echo T1-weighted, gradient recalled echo chemical-shift T1-weighted, and contrast-enhanced gradient recalled echo T1-weighted sequences. POPULATION: One hundred five patients with ccRCC who underwent preoperative contrast-enhanced MRI and subsequent genomic sequencing: 59 consecutive patients from our institution (38 [64.41%] with VHL mutations) formed a training cohort, and 46 from The Cancer Genome Atlas (TCGA) database (24 [52.17%] with VHL mutations) formed an independent test cohort. ASSESSMENT: Two radiologists, with 23 and 33 years of experience respectively, jointly evaluated the semantic MRI features of the primary lesion in ccRCCs to propose potential features related to VHL mutations in both cohorts. Three additional readers, with 5, 7, and 10 years of experience respectively, independently reviewed all lesions to assess the interobserver agreement of MRI features. A VHL mutational likelihood score (VHL-MULIS) system was constructed using the training cohort and validated using the independent test cohort. STATISTICAL TESTS: Fisher's test or chi-square test, t-test or Mann-Whitney U test, logistic regression, Cohen's kappa (κ), area under the receiver operating characteristic curve (AUC). A two-sided P value <0.05 was considered statistically significant. RESULTS: In both the local and public cohorts, T2-weighted signal intensity and presence of microscopic fat from primary lesions were significantly associated with VHL mutation status. The VHL-MULIS incorporated maximum diameter, T2-weighted signal intensity, and presence of microscopic fat in the training cohort and demonstrated promising diagnostic ability (AUC, 0.82; sensitivity, 0.79; specificity, 0.82) and substantial interobserver agreement (κ, 0.787) in the test cohort. DATA CONCLUSION: The VHL mutation exhibited a distinct MRI phenotype. Integrating multiple semantic MRI features has potential to reflect the mutation status in patients with ccRCC. EVIDENCE LEVEL: 3 TECHNICAL EFFICACY: Stage 2.

Evaluation of Metabolic Engineering Strategies on 2-Ketoisovalerate Production by Escherichia coli.

As an important metabolic intermediate, 2-ketoisovalerate has significant potential in the pharmaceutical and biofuel industries. However, a low output through microbial fermentation inhibits its industrial application. The microbial production of 2-ketoisovalerate is representative whereby redox imbalance is generated with two molecules of NADH accumulated and an extra NADPH required to produce one 2-ketoisovalerate from glucose. To achieve efficient 2-ketoisovalerate production, metabolic engineering strategies were evaluated in Escherichia coli. After deleting the competing routes, overexpressing the key enzymes for 2-ketoisovalerate production, tuning the supply of NADPH, and recycling the excess NADH through enhancing aerobic respiration, a 2-ketoisovalerate titer and yield of 46.4 g/L and 0.644 mol/mol glucose, respectively, were achieved. To reduce the main by-product of isobutanol, the activity and expression of acetolactate synthase were modified. Additionally, a protein degradation tag was fused to pyruvate dehydrogenase (PDH) to curtail the conversion of pyruvate precursor into acetyl-CoA and the generation of NADH. The resulting strain, 050TY/pCTSDTQ487S-RBS55, was initially incubated under aerobic conditions to attain sufficient cell mass and then transferred to a microaerobic condition to degrade PDH and inhibit the remaining activity of PDH. Intracellular redox imbalance was relieved with titer, productivity and yield of 2-ketoisovalerate improved to 55.8 g/L, 2.14 g/L h and 0.852 mol/mol glucose. These results revealed metabolic engineering strategies for the production of a redox-imbalanced fermentative metabolite with high titer, productivity, and yield. IMPORTANCE An efficient microbial strain was constructed for 2-ketoisovalerate synthesis. The positive effect of the leuA deletion on 2-ketoisovalerate production was found. An optimal combination of overexpressing the target genes was obtained by adjusting the positions of the multiple enzymes on the plasmid frame and the presence of terminators, which could also be useful for the production of downstream products such as isobutanol and l-valine. Reducing the isobutanol by-product by engineering the acetolactate synthase called for special attention to decreasing the promiscuous activity of the enzymes involved. Redox-balancing strategies such as tuning the expression of the chromosomal pyridine nucleotide transhydrogenase, recycling NADH under aerobic cultivation, switching off PDH by degradation, and inhibiting the expression and activity under microaerobic conditions were proven effective for improving 2-ketoisovalerate production. The degradation of PDH and inhibiting this enzyme's expression would serve as a means to generate a wide range of products from pyruvate.

Rapid Mining of Novel α-Glucosidase and Lipase Inhibitors from Streptomyces sp. HO1518 Using UPLC-QTOF-MS/MS.

A rapid and sensitive method using ultra-high performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS/MS) was applied for the analysis of the metabolic profile of acarviostatin-containing aminooligosaccharides derived from Streptomyces sp. HO1518. A total of ninety-eight aminooligosaccharides, including eighty potential new compounds, were detected mainly based on the characteristic fragment ions originating from quinovosidic bond cleavages in their molecules. Following an LC-MS-guided separation technique, seven new aminooligosaccharides (10-16) along with four known related compounds (17-20) were obtained directly from the crude extract of strain HO1518. Compounds 10-13 represent the first examples of aminooligosaccharides with a rare acarviostatin II02-type structure. In addition, all isolates displayed considerable inhibitory effects on three digestive enzymes, which revealed that the number of the pseudo-trisaccharide core(s), the feasible length of the oligosaccharides, and acyl side chain exerted a crucial influence on their bioactivities. These results demonstrated that the UPLC-QTOF-MS/MS-based metabolomics approach could be applied for the rapid identification of aminooligosaccharides and other similar structures in complex samples. Furthermore, this study highlights the potential of acylated aminooligosaccharides with conspicuous α-glucosidase and lipase inhibition for the future development of multi-target anti-diabetic drugs.

Grazing intensity changed the activities of nitrogen assimilation related enzymes in desert Steppe Plants.

BACKGROUND: Nitrogen, as a limiting factor for net primary productivity in grassland ecosystems, is an important link in material cycles in grassland ecosystems. However, the nitrogen assimilation efficiency and mechanisms of grassland plants under grazing disturbance are still unclear. This study investigated Stipa breviflora desert steppe which had been grazed for 17 years and sampled the root system and leaf of the constructive species Stipa breviflora during the peak growing season under no grazing, light grazing, moderate grazing and heavy grazing treatments. The activities of enzymes related to nitrogen assimilation in roots and leaves were measured. RESULTS: Compared with no grazing, light grazing and moderate grazing significantly increased the activities of nitrate reductase (NR), glutamine synthetase (GS), glutamic oxaloacetic transaminase (GOT) and glutamic pyruvate transaminase (GPT) in leaves, and GS, GOT and GPT in roots of Stipa breviflora, while heavy grazing significantly decreased the activities of GS in leaves and NR in roots of Stipa breviflora. NR, GOT and GPT activities in leaves and roots of Stipa breviflora were positively correlated with nitrogen content, soluble protein, free amino acid and nitrate content. CONCLUSIONS: Grazing disturbance changed the activities of nitrogen assimilation related enzymes of grassland plants, and emphasized that light grazing and moderate grazing were beneficial for nitrogen assimilation by grassland plants. Therefore, establishing appropriate stocking rates is of great significance for material flows in this grassland ecosystem and for the stability and sustainable utilization of grassland resources.

Engineering the Outer Membrane Could Facilitate Better Bacterial Performance and Effectively Enhance Poly-3-Hydroxybutyrate Accumulation.

Poly-3-hydroxybutyrate (PHB) is an environmentally friendly polymer and can be produced in Escherichia coli cells after overexpression of the heterologous gene cluster phaCAB. The biosynthesis of the outer membrane (OM) consumes many nutrients and influences cell morphology. Here, we engineered the OM by disrupting all gene clusters relevant to the polysaccharide portion of lipopolysaccharide (LPS), colanic acid (CA), flagella, and/or fimbria in E. coli W3110. All these disruptions benefited PHB production. Especially, disrupting all these OM components increased the PHB content to 83.0 wt% (PHB content percentage of dry cell weight), while the wild-type control produced only 1.5 wt% PHB. The increase was mainly due to the LPS truncation to Kdo2 (3-deoxy-d-manno-octulosonic acid)-lipid A, which resulted in 82.0 wt% PHB with a 25-fold larger cell volume, and disrupting CA resulted in 57.8 wt% PHB. In addition, disrupting LPS facilitated advantageous fermentation features, including 69.1% less acetate, a 550% higher percentage of autoaggregated cells among the total culture cells, 69.1% less biofilm, and a higher broken cell ratio. Further detailed mechanism investigations showed that disrupting LPS caused global changes in envelope and cellular metabolism: (i) a sharp decrease in flagella, fimbria, and secretions; (ii) more elastic cells; (iii) much greater carbon flux toward acetyl coenzyme A (acetyl-CoA) and supply of cofactors, including NADP, NAD, and ATP; and (iv) a decrease in by-product acids but increase in γ-aminobutyric acid by activating σE factor. Disrupting CA, flagella, and fimbria also improved the levels of acetyl-CoA and cofactors. The results indicate that engineering the OM is an effective strategy to enhance PHB production and highlight the applicability of OM engineering to increase microbial cell factory performance. IMPORTANCE Understanding the detailed influence of the OM on the cell envelope and cellular metabolism is important for optimizing the E. coli cell factory and many other microorganisms. This study revealed the applicability of remodeling the OM to enhance PHB accumulation as representative inclusion bodies. The results generated in this study give essential information for producing other inclusion bodies or chemicals which need more acetyl-CoA and cofactors but less by-product acids. This study is promising to provide new ideas for the improvement of microbial cell factories.

Microbial Oligosaccharides with Biomedical Applications.

Microbial oligosaccharides have been regarded as one of the most appealing natural products attributable to their potent and selective bioactivities, such as antimicrobial activity, inhibition of α-glucosidases and lipase, interference of cellular recognition and signal transduction, and disruption of cell wall biosynthesis. Accordingly, a handful of bioactive oligosaccharides have been developed for the treatment of bacterial infections and type II diabetes mellitus. Given that naturally occurring oligosaccharides have increasingly gained recognition in recent years, a comprehensive review is needed. The current review highlights the chemical structures, biological activities and divergent biosynthetic origins of three subgroups of oligomers including the acarviosine-containing oligosaccharides, saccharomicins, and orthosomycins.

Acylated Aminooligosaccharides from the Yellow Sea Streptomyces sp. HO1518 as Both α-Glucosidase and Lipase Inhibitors.

Three new acylated aminooligosaccharide (1-3), along with five known congeners (4-8), were isolated from the marine-derived Streptomyces sp. HO1518. Their structures were fully elucidated by extensive spectroscopic analysis, mainly based on 1D-selective and 2D TOCSY, HSQC-TOCSY, and HRESIMS spectrometry measurements, and by chemical transformations. All of the compounds were evaluated for their α-glucosidase and pancreatic lipase inhibitory activities. Among the isolates, D6-O-isobutyryl-acarviostatin II03 (3) and D6-O-acetyl-acarviostatin II03 (8), sharing acarviostatin II03-type structure, showed the most potent α-glucosidase and lipase inhibitory effects, far stronger than the antidiabetic acarbose towards α-glucosidase and almost equal to the anti-obesity orlistat towards lipase in vitro. This is the first report on inhibitory activities against the two major digestive enzymes for acylated aminooligosaccharides. The results from our investigation highlight the potential of acylated aminooligosaccharides for the future development of multi-target anti-diabetic drug.

A new brominated polyacetylene from Chinese marine sponge Xestospongia testudinaria.

A new brominated polyacetylene xestonariene J (1), bearing a rare 2,4-dibromo-1-ene-3-yne terminal in the molecule, along with two known related analogues (2 and 3), was obtained from Chinese marine sponge Xestospongia testudinaria. Its structure was determined on the basis of detailed spectroscopic analysis and comparison with literature data.

Gene-Wide Analysis of Aquaporin Gene Family in Malus domestica and Heterologous Expression of the Gene MpPIP2;1 Confers Drought and Salinity Tolerance in Arabidposis thaliana.

The aquaporins (AQPs) are a family of integral membrane proteins involved in the transcellular membrane transport of water and other small molecules. A scan of the apple (Malus domestica) genome revealed the presence of 42 genes encoding putative AQPs. Based on a phylogenetic analysis of the deduced peptide sequences of the AQPs generated by Arabidopsis thaliana, poplar (Populus trichocarpa), and rubber (Hevea brasiliensis), the apple AQPs were each assigned membership of the five established AQP subfamilies, namely the PIPs (eleven members), the TIPs (thirteen members), the NIPs (eleven members), the SIPs (five members), and the XIPs (two members). The apple AQPs included asparagine-proline-alanine (NPA) motifs, an aromatic/arginine (ar/R) selectivity filter, and the Froger's positions. The heterologous expression of MpPIP2;1 in A. thaliana was shown to enhance the level of tolerance exhibited against both drought and salinity.

Acylated Aminooligosaccharides with Inhibitory Effects against α-Amylase from Streptomyces sp. HO1518.

Five new acylated aminooligosaccharides (1⁻5), together with one known related analogue (6), were isolated from Streptomyces sp. HO1518. Their structure was identified by extensive spectroscopic analysis, including 1D and 2D NMR data and high resolution electrospray ionization mass spectrometry (HRESIMS), and by comparison with those reported in the literature. All of the new compounds showed more promising porcine pancreatic α-amylase (PPA) inhibitory activities than the clinical drug acarbose, indicating them as potential pharmaceutical drug leads toward type II diabetes.

Enhancement of ε-poly-L-lysine synthesis in Streptomyces by exogenous glutathione.

Our previous work indicated that the vigor of Streptomyces decreased at the later stage of ε-poly-L-lysine (ε-PL) fermentation. In this study, we observed that the level of reactive oxygen species (ROS) in vivo increased sharply after 24 h, and the addition of an antioxidant glutathione (GSH) before this increase in ROS stimulated ε-PL synthesis in shake-flask fermentation. The enhancement of ε-PL production by GSH was further verified in batch and fed-batch fermentations. On a 5-l fermenter scale, the highest increasement was 68.8% in batch fermentation and the highest ε-PL level was 46.5 g l- 1 in fed-batch fermentation. The RT-qPCR analysis showed that the transcriptional level of the catalase gene was down-regulated, and the decrease in cell activity was alleviated by the addition of GSH. The results revealed that exogenous antioxidant might maintain the cell vigor by reducing the excess ROS which provided a novel approach to regulate ε-PL synthesis.

Further brominated polyacetylenes with pancreatic lipase inhibitory activity from Chinese marine sponge Xestospongia testudinaria.

A new brominated polyacetylene, xestonariene I (1), along with three known related analogues (2-4), was obtained from Chinese marine sponge Xestospongia testudinaria. Its structure was determined on the basis of detailed spectroscopic analysis and by comparison with literature data. Compound 4 exhibited significant inhibitory activity against pancreatic lipase, which plays a key role in preventing obesity, with an IC50 value of 0.61 µM, being comparable to that of the positive control orlistat (IC50 = 0.78 µM).

Aggregation-induced emission and intermolecular charge transfer effect in triphenylamine fluorophores containing diphenylhydrazone structures.

Three new chromophores incorporating acceptor-π-donor-π-acceptor structural motifs and mono-, di- and tri-branched diphenylsulfone base linked to triphenylamine through a hydrazone π-bridge were synthesized, and the photoluminescence properties of the three chromophores were studied in solutions as well as in aggregated states. All the fluorophores emitted strong blue fluorescence in THF. Mono- and di-branched triphenylamine both exhibited increasing blue fluorescence and displayed an AIEE effect in the aggregated state. Tri-branched triphenylamine emitted green fluorescence and presented the AIE effect in the aggregated state. These interesting phenomena have been interpreted by a molecular stacking mode with molecular dynamics (MD) and DFT calculations. The unique propeller shaped molecular configuration of triphenylamine prevented face to face π-π stacking and induced the hindered rotation, which resulted in the AIEE or AIE effect in the aggregated state. The enlarged coplanarity of diphenylhydrazone chains increased the conjugation of tri-branched triphenylamine, which was beneficial to the formation of ICT and AIE and resulted in emitting green ICT fluorescence in the aggregated state. Fluorescent microscope imaging and the fluorescent pictures of the powder states certified the strong AIEE effect or AIE effect in the solid.

A new bioactive steroidal ketone from the South China Sea sponge Xestospongia testudinaria.

A new steroidal ketone (1), with an ergosta-22,25-diene side chain, was obtained from the South China Sea marine sponge Xestospongia testudinaria. The structure of 1 was determined on the basis of detailed spectroscopic analysis and by comparison with literature. Compound 1 exhibited significant inhibitory activity against protein tyrosine phosphatase 1B (PTP1B), a key target for the treatment of type II diabetes and obesity, with an IC50 value of 4.27 ± 0.55 µM, which is comparable with the positive control oleanolic acid (IC50 = 2.63 ± 0.22 µM).

Brominated polyunsaturated lipids with protein tyrosine phosphatase-1B inhibitory activity from Chinese marine sponge Xestospongia testudinaria.

A new brominated polyunsaturated lipid, methyl (E,E)-14,14-dibromo-4,6,13-tetradecatrienoate (1), along with three known related analogues (2-4), were isolated from the Et2O-soluble portion of the acetone extract of Chinese marine sponge Xestospongia testudinaria treated with diazomethane. The structure of the new compound was elucidated by detailed spectroscopic analysis and by comparison with literature data. Compound 3 exhibited significant inhibitory activity against protein tyrosine phosphatase 1B (PTP1B), a key target for the treatment of type II diabetes and obesity, with an IC50 value of 5.30 ± 0.61 µM, when compared to the positive control oleanolic acid (IC50 = 2.39 ± 0.26 µM).

Experimental evaluation of myocardial fibrosis in a rapid atrial pacing model in New Zealand rabbits using quantitative analysis of ultrasonic backscatter.

BACKGROUND: The aim of this study was the establishment of a rapid atrial pacing (RAP)-induced atrial fibrillation (AF) model with electrophotoluminescence and the application of ultrasonic backscatter quantitative analysis of the degree of myocardial fibrosis in New Zealand white rabbits. MATERIAL AND METHODS: Sixteen New Zealand white rabbits were randomly divided into 2 groups: 1) a sham operation group (n=8) with implanted electrodes and no rapid pacing and 2) a pacing group (n=8) with an AF model induced by short-term rapid right atrial pacing for 12 h. Establishment of an AF model, atrial myocardium of myocardial fibrosis was tested by Masson staining and expression of collagen I and collagen III protein was detected with pathologic immunohistochemistry integrated back-scatter (IBS). Back scattering integral cycle variation (CVIB) were detected in atrial septal and posterior wall of the right atrium. RESULTS: Rapid atrial pacing successfully induced the atrial fibrillation model in rabbits. Masson staining showed myocardial fibrosis significantly increased in the pacing group. Expression of collagen I and collagen III protein was strongly positive in the pacing group, and expression of collagen I and collagen III protein were weakly positive in the sham operation group. Compared with the sham operation group, AII was increased (8.24±0.85 vs. 15.56±1.30, P<0.05) and (7.58±0.56 vs. 16.60±2.45, P<0.05). CVIB was significantly decreased (2.78±0.86 vs. 1.08±0.13, P<0.05) and (3.12±0.65 vs. 1.56±0.15, P<0.05) in septal and posterior wall of the right atrium of the pacing group. CONCLUSIONS: Ultrasonic backscatter measurement technique can be used to evaluate degree of myocardial fibrosis in a right atrial pacing-induced atrial arrhythmia model.

Four phragmalin orthoesters from the Chinese mangrove Xylocarpus granatum.

Four new 8,9,30-phragmalin orthoesters (1-4), along with six related known compounds, namely xyloccensins O-S (5-9) and V (10), were isolated and characterized from the twigs and leaves of the Chinese mangrove Xylocarpus granatum. The structures of the new compounds were determined on the basis of extensive spectroscopic analysis and by comparison with those of related known compounds in the literature. The absolute configuration of xyloccensin Q (7) was revised as its enantiomer by X-ray diffraction analysis employing graphite monochromated Cu Kα radiation (λ=1.54178 Å) with a Flack parameter of -0.04 and was further secured by a time-dependent density functional theory electronic circular dichroism (TDDFT ECD) calculation. Consequently, the absolute configurations of xyloccensins O (5), P (6), R (8), S (9), and V (10) were all corrected as their corresponding enantiomers, respectively. Xyloccensin S (9) exhibited inhibitory activity against protein tyrosine phosphatase 1B, a potential drug target for the treatment of type II diabetes and obesity, with an IC50 value of 8.72 µg/mL.

Thunberol, a new sterol from the Chinese brown alga Sargassum thunbergii.

A new sterol, named thunberol (1), along with four known analogs, 24-ethylcholesta-4,24(28)-dien-3-one (2), stigmasta-5,28-dien-3ß-ol (3), cholesta-5,14-dien-3ß-ol (4), and cholesta-5,23-dien-3ß,25-diol (5), were isolated from the brown alga Sargassum thunbergii collected from East China Sea. The structures of these metabolites were elucidated on the basis of detailed spectroscopic analysis and by comparison with the literature data. Thunberol (1) exhibited significant inhibitory activity against protein tyrosine phosphatase 1B, a potential drug target for the treatment of Type-II diabetes and obesity, with an IC50 value of 2.24 µg/ml.

[Review on the secondary metabolites from Xestospongia sponges and their bioactivities].

The genus Xestospongia is one of the most widespread genera of sponges, containing abundant secondary metatolites with novel structures and potent bioactivities. The main structure types of secondary metatolites found in this genus are alkaloids, quinines, terpens, steroids, lipids, polyketones, etc. These metatolites exhibit a variety of bioactivities, such as cytotoxic, antibacterial and antiviral activities. This paper reviews the progress in the chemistry and pharmacological activities of the second metabolities from sponges of Xestospongia, especially for recent five years, with the aim for further research.