Observation of Subdiffusive Dynamic Scaling in a Driven and Disordered Bose Gas.

We explore the dynamics of a tuneable box-trapped Bose gas under strong periodic forcing in the presence of weak disorder. In absence of interparticle interactions, the interplay of the drive and disorder results in an isotropic nonthermal momentum distribution that shows subdiffusive dynamic scaling, with sublinear energy growth and the universal scaling function captured well by a compressed exponential. We explain that this subdiffusion in momentum space can naturally be understood as a random walk in energy space. We also experimentally show that for increasing interaction strength, the gas behavior smoothly crosses over to wave turbulence characterized by a power-law momentum distribution, which opens new possibilities for systematic studies of the interplay of disorder and interactions in driven quantum systems.

Serum metabolomics analysis of biomarkers and metabolic pathways in patients with colorectal cancer associated with spleen-deficiency and qi-stagnation syndrome or damp-heat syndrome: a prospective cohort study.

Objective: To profile the serum metabolites and metabolic pathways in colorectal cancer (CRC) patients associated with spleen-deficiency and qi-stagnation syndrome (SDQSS) or damp-heat syndrome (DHS). Methods: From May 2020 to January 2021, CRC patients diagnosed with traditional Chinese medicine (TCM) syndromes of SDQSS or DHS were enrolled. The clinicopathological data of the SDQSS and DHS groups were compared. The serum samples were analyzed by liquid chromatography-mass spectrometry (LC-MS). The variable importance in the projection >1, fold change ≥3 or ≤0.333, and P value ≤0.05 were used to identify differential metabolites between the two groups. Furthermore, areas under the receiver operating characteristic (ROC) curve > 0.9 were applied to select biomarkers with good predictive performance. The enrichment metabolic pathways were searched through the database of Kyoto Encyclopedia of Genes and Genomes. Results: 60 CRC patients were included (30 SDQSS and 30 DHS). The level of alanine aminotransferase was marginally significantly higher in the DHS group than the SDQSS group (P = 0.051). The other baseline clinicopathological characteristics were all comparable between the two groups. 23 differential serum metabolites were identified, among which 16 were significantly up-regulated and 7 were significantly down-regulated in the SDQSS group compared with the DHS group. ROC curve analysis showed that (S)-3-methyl-2-oxopentanoic acid, neocembrene, 1-aminocyclopropanecarboxylic acid, 3-methyl-3-hydroxypentanedioate, and nicotine were symbolic differential metabolites with higher predictive power. The top five enrichment signalling pathways were valine, leucine and isoleucine biosynthesis; lysosome; nicotine addiction; fructose and mannose metabolism; and pertussis. Conclusion: Our study identifies the differential metabolites and characteristic metabolic pathways among CRC patients with SDQSS or DHS, offering the possibility of accurate and objective syndrome differentiation and TCM treatment for CRC patients.

Genome-wide identification and expression pattern analysis of R2R3-MYB transcription factor gene family involved in puerarin biosynthesis and response to hormone in Pueraria lobata var. thomsonii.

BACKGROUND: R2R3-MYB transcription factors regulate secondary metabolism, stress responses and development in various plants. Puerarin is a bioactive ingredient and most abundant secondary metabolite isolated from Pueraria lobata. The biosynthesis of puerarin proceeds via the phenylpropanoid pathway and isoflavonoids pathway, in which 9 key enzymes are involved. The expression of these structural genes is under control of specific PtR2R3-MYB genes in different plant tissues. However, how PtR2R3-MYB genes regulates structural genes in puerarin biosynthesis remains elusive. This study mined the PtR2R3-MYB genes involved in puerarin biosynthesis and response to hormone in Pueraria lobata var. thomsonii. RESULTS: A total of 209 PtR2R3-MYB proteins were identified, in which classified into 34 subgroups based on the phylogenetic topology and the classification of the R2R3-MYB superfamily in Arabidopsis thaliana. Furtherly physical and chemical characteristics, gene structure, and conserved motif analysis were also used to further analyze PtR2R3-MYBs. Combining puerarin content and RNA-seq data, speculated on the regulated puerarin biosynthesis of PtR2R3-MYB genes and structural genes, thus 21 PtR2R3-MYB genes and 25 structural genes were selected for validation gene expression and further explore its response to MeJA and GSH treatment by using qRT-PCR analysis technique. Correlation analysis and cis-acting element analysis revealed that 6 PtR2R3-MYB genes (PtMYB039, PtMYB057, PtMYB080, PtMYB109, PtMYB115 and PtMYB138) and 7 structural genes (PtHID2, PtHID9, PtIFS3, PtUGT069, PtUGT188, PtUGT286 and PtUGT297) were directly or indirectly regulation of puerarin biosynthesis in ZG11. It is worth noting that after MeJA and GSH treatment for 12-24 h, the expression changes of most candidate genes were consistent with the correlation of puerarin biosynthesis, which also shows that MeJA and GSH have the potential to mediate puerarin biosynthesis by regulating gene expression in ZG11. CONCLUSIONS: Overall, this study provides a comprehensive understanding of the PtR2R3-MYB and will paves the way to reveal the transcriptional regulation of puerarin biosynthesis and response to phytohormone of PtR2R3-MYB genes in Pueraria lobata var. thomsonii.

Molecular Cloning and Functional Characterization of a Sterol 3-O-Glucosyltransferase Involved in Biosynthesis of Steroidal Saponins in Trigonella foenum-graecum.

Fenugreek (Trigonella foenum-graecum), a pharmacologically important herb, is widely known for its antidiabetic, hypolipidemic, and anticancer effects. The medicinal properties of this herb are accredited to the presence of bioactive steroidal saponins with one or more sugar moieties linked to the C-3 OH position of disogenin or its C25-epimer yamogenin. Despite intensive studies regarding pharmacology and phytochemical profiles of this plant, enzymes and/or genes involved in synthesizing the glycosidic part of fenugreek steroidal saponins are still missing so far. This study reports the molecular cloning and functional characterization of a key sterol-specific glucosyltransferase, designated as TfS3GT2 here, from fenugreek plant. The recombinant TfS3GT2 was purified via expression in Escherichia coli, and biochemical characterization of the recombinant enzyme suggested its role in transferring a glucose group onto the C-3 hydroxyl group of diosgenin or yamogenin. The functional role of TfS3GT2 in the steroidal saponin biosynthesis was also demonstrated by suppressing the gene in the transgenic fenugreek hairy roots via the RNA interference (RNAi) approach. Down-regulation of TfS3GT2 in fenugreek generally led to reduced levels of diosgenin or yamogenin-derived steroidal saponins. Thus, Tf3SGT2 was identified as a steroid-specific UDP-glucose 3-O-glucosyltransferase that appears to be involved in steroidal saponin biosynthesis in T. foenum-graecum.

Engineering of Saccharomyces cerevisiae for 24-Methylene-Cholesterol Production.

24-Methylene-cholesterol is a necessary substrate for the biosynthesis of physalin and withanolide, which show promising anticancer activities. It is difficult and costly to prepare 24-methylene-cholesterol via total chemical synthesis. In this study, we engineered the biosynthesis of 24-methylene-cholesterol in Saccharomyces cerevisiae by disrupting the two enzymes (i.e., ERG4 and ERG5) in the yeast's native ergosterol pathway, with ERG5 being replaced with the DHCR7 (7-dehydrocholesterol reductase) enzyme. Three versions of DHCR7 originating from different organisms-including the DHCR7 from Physalis angulata (PhDHCR7) newly discovered in this study, as well as the previously reported OsDHCR7 from Oryza sativa and XlDHCR7 from Xenopus laevis-were assessed for their ability to produce 24-methylene-cholesterol. XlDHCR7 showed the best performance, producing 178 mg/L of 24-methylene-cholesterol via flask-shake cultivation. The yield could be increased up to 225 mg/L, when one additional copy of the XlDHCR7 expression cassette was integrated into the yeast genome. The 24-methylene-cholesterol-producing strain obtained in this study could serve as a platform for characterizing the downstream enzymes involved in the biosynthesis of physalin or withanolide, given that 24-methylene-cholesterol is a common precursor of these chemicals.

De Novo Biosynthesis of the Anticancer Compound Euphol in Saccharomyces cerevisiae.

Euphol is a euphane-type tetracyclic triterpene which is primarily found in the Euphorbia genus. Euphol has been renowned because of its great potential as a promising anticancer drug. Surprisingly, despite its diverse antitumor effects, the respective gene for euphol biosynthesis had not been identified until this study. In our experiments with Euphorbia tirucalli, euphol was detected predominantly in latex, the element that is often used for cancer treatments in Brazil. Two latex-specifically expressed oxidosqualene cyclases (OSCs) from E. tirucalli, designated as EtOSC5 and EtOSC6, were functionally characterized by expression in a lanosterol synthase knockout yeast strain GIL77. EtOSC5 produces euphol and its 20S-isomer tirucallol as two of the major products, while EtOSC6 produces taraxasterol and ß-amyrin as the major products. These four compounds were also detected as the major triterpenes in the E. tirucalli latex, suggesting that EtOSC5 and EtOSC6 are the primary catalysts for the formation of E. tirucalli latex triterpene alcohols. Based on a model structure of EtOSC5 followed with site-mutagenesis experiments, the mechanism for the EtOSC5 activity was proposed. By applying state-of-the-art engineering techniques, the expression of EtOSC5 together with three other known precursor genes were chromosomally integrated into Saccharomyces cerevisiae. The resulting engineered yeast strain YS5E-1 produced 1.84 ± 0.17 mg/L of euphol in shake flasks.

Identification and Characterization of a Trillin Rhamnosyltransferase From Dioscorea zingiberensis.

Dioscorea zingiberensis accumulates abundant steroidal saponins, such as dioscin, which is the principal bioactive ingredient displaying a wide range of pharmacological activities. Diosgenin is the aglycone of dioscin, and recently, genes encoding cytochrome P450 enzymes in the late steps of diosgenin biosynthesis have been isolated. Diosgenin was successfully synthesized in the cholesterol-producing yeasts. From diosgenin to dioscin, one glucose and two rhamnose groups need to be added. Although genes encoding UDP-glucosyltransferases converting diosgenin to trillin were isolated, genes encoding UDP-rhamnosyltransferases involved in dioscin biosynthesis remain unknown. In this study, we isolated the cDNA encoding the trillin rhamnosyltransferase (designated DzGT1) from D. zingiberensis. Heterologous expression of DzGT1 in Escherichia coli cells showed that the gene product exhibits an enzyme activity that glycosylates the trillin to form prosapogenin A of dioscin (PSA). The transcript level of DzGT1 is in accord with PSA accumulation in different organs of D. zingiberensis. Integration of the biochemical, metabolic, and transcriptional data supported the function of DzGT1 in dioscin biosynthesis. The identification and characterization of DzGT1 will help understand the metabolism of steroidal saponins in D. zingiberensis and provide candidate UDP-rhamnosyltransferase for efficient production of PSA, dioscin, and relevant steroidal saponins in microbial hosts.

Feedback of biochemical indexes of blood sugar control by swimming exercise / Feedback dos índices bioquímicos de controle de açúcar no sangue por exercícios de natação / Feedback de los índices bioquímicos del control del azúcar en sangre mediante el ejercicio de natación

ABSTRACT Introduction: According to the metabolic characteristics of ultra-long-distance swimming and the characteristics of energy utilization and absorption during exercise, we have formulated a nutritional supplement plan for crossing to study the influence of swimming sports on blood sugar and give biochemical feedback indicators. Objective: To lay a foundation for studying the nutrition supplement rules during long-term exercise by taking the athletes' blood after training to determine the changes in blood sugar, adjusting and determining the nutritional supplement plan during training. Methods: We monitor athletes' physical function changes and biochemical indicators during training and study the changes of these biochemical indicators and athletes' physical functions after long-term swimming exercises to scientifically arrange the exercise intensity and load during the training period. Results: The urine indexes after exercise did not change much, reflecting the exercise load's low intensity. The changes in blood lactic acid and blood urea indexes after exercise also confirmed this. During the training period, the athletes' hemoglobin and red blood cell parameters are in the ideal range, indicating that the athlete's physical function is in good condition. During the training period, the training load intensity and load are reasonable according to ultra-long-distance swimming's energy supply characteristics. After training, the changes in blood glucose indicators reflect that the nutritional supplement program we formulated for athletes is reasonable and feasible. Conclusions: By monitoring the blood sugar and biochemical indicators of swimmers, it can help athletes to arrange exercise intensity scientifically and load during the training period, to better carry out open water competitions in China, and to arrange training and scientific nutrition during the training period scientifically. Lay the foundation for the establishment of nutrition supplement theory and training theory for super long-time sports. Level of evidence II; Therapeutic studies - investigation of treatment results.

RESUMEN Introducción: De acuerdo con las características metabólicas de la natación de ultra larga distancia y las características de utilización y absorción de energía durante el ejercicio, hemos formulado un plan de complementos nutricionales para estudiar la influencia de los deportes de natación en el azúcar en sangre y dar indicadores de retroalimentación bioquímica. Objetivo: Sentar las bases para el estudio de las reglas de los suplementos nutricionales durante el ejercicio a largo plazo mediante la extracción de sangre de los atletas después del entrenamiento para determinar los cambios en el azúcar en sangre, ajustando y determinando el plan de suplementos nutricionales durante el entrenamiento. Métodos: monitoreamos los cambios en la función física de los atletas y los indicadores bioquímicos durante el entrenamiento y estudiamos los cambios de estos indicadores bioquímicos y las funciones físicas de los atletas después de ejercicios de natación de larga distancia para organizar científicamente la intensidad y la carga del ejercicio durante el período de entrenamiento. Resultados: Los índices de orina después del ejercicio no cambiaron mucho, lo que refleja la baja intensidad de la carga de ejercicio. Los cambios en los índices de ácido láctico y urea en sangre después del ejercicio también lo confirmaron. Durante el período de entrenamiento, los parámetros de hemoglobina y glóbulos rojos de los atletas están en el rango ideal, lo que indica que la función física del atleta está en buenas condiciones. Durante el período de entrenamiento, la intensidad de la carga de entrenamiento y la carga son razonables de acuerdo con las características de suministro de energía de la natación de ultra larga distancia. Después del entrenamiento, los cambios en los indicadores de glucosa en sangre reflejan que el programa de suplementos nutricionales que formulamos para los atletas es razonable y factible. Conclusiones: monitorear los indicadores bioquímicos y de azúcar en sangre de los nadadores, puede ayudar a los atletas a organizar científicamente la intensidad del ejercicio y la carga durante el período de entrenamiento, a realizar mejor las competiciones en aguas abiertas en China y a organizar el entrenamiento y la nutrición científica durante el período de entrenamiento. Sentar las bases para el establecimiento de la teoría de los suplementos nutricionales y la teoría del entrenamiento para deportes de larga duración. Nivel de evidencia II; Estudios terapéuticos: investigación de los resultados del tratamiento.

RESUMO Introdução: De acordo com as características metabólicas da natação de ultra longa distância e as características de utilização e absorção de energia durante o exercício, formulamos um plano de suplemento nutricional para estudar a influência dos esportes de natação no açúcar no sangue e fornecer indicadores de feedback bioquímico. Objetivo: Estabelecer as bases para o estudo das regras de suplementos nutricionais durante exercícios de longa duração, retirando sangue de atletas após o treinamento para determinar as mudanças na glicemia, ajustando e determinando o plano de suplementação nutricional durante o treinamento. Métodos: monitoramos as mudanças na função física e nos indicadores bioquímicos dos atletas durante o treinamento e estudamos as mudanças nesses indicadores bioquímicos e nas funções físicas dos atletas após exercícios de natação de longa distância para organizar cientificamente a intensidade e a carga do exercício durante o período de treinamento. Resultados: As taxas de urina após o exercício não mudaram muito, refletindo a baixa intensidade da carga de exercício. Alterações nos índices de uréia e ácido láctico no sangue após o exercício também confirmaram isso. Durante o período de treinamento, os parâmetros de hemoglobina e hemácias dos atletas estão na faixa ideal, indicando que a função física do atleta está em boas condições. Durante o período de treinamento, a intensidade da carga de treinamento e a carga são razoáveis de acordo com as características da fonte de alimentação da natação de ultra longa distância. Após o treinamento, as mudanças nos indicadores de glicose no sangue refletem que o programa de suplementos nutricionais que formulamos para atletas é razoável e viável. Conclusões: monitorar os indicadores bioquímicos e de açúcar no sangue de nadadores pode ajudar os atletas a organizar cientificamente a intensidade e carga do exercício durante o período de treinamento, conduzir melhor competições em águas abertas na China e organizar treinamento e nutrição científica durante o período de treinamento. Estabelecendo as bases para o estabelecimento da teoria dos suplementos nutricionais e da teoria do treinamento para esportes de longa duração. Nível de evidência II; Estudos terapêuticos: investigação dos resultados do tratamento.

Growth of gas-filled penny-shaped cracks in decompressed hydrogels.

We report that the decompression of soft brittle materials can lead to the growth of internal gas-filled cracks. These cracks are oblate spheroids ('penny shape'), whose major radius grows linearly in time, irreversibly fracturing the surrounding material. Our optical measurements in hydrogels characterise and quantify the three-dimensional crack geometry and growth rate. These results are in good agreement with our analytical model coupling fracture mechanics and gas diffusion, and predicting the dependence on the mechanical properties, gas diffusivity and super-saturation conditions (gas pressure, solubility, temperature). Our results suggest a new potential mechanism for decompression sickness in scuba diving and for indirect optical measurements of the fracture properties of hydrogels.

[Expression of CD44v6 and COX-2 in pleomorphic adenomas and carcinomas of parotid gland].

PURPOSE: To investigate the correlation between CD44v6 and cyclooxygenase-2(COX-2) expression and recurrence and deterioration of pleomorphic adenoma of parotid gland. METHODS: Immunohistochemistry was used to detect the expression of CD44v6 and COX-2 in 19 normal parotid tissues, 25 primary pleomorphic adenomas, 15 first recurrent pleomorphic adenoma, 15 second recurrent pleomorphic adenoma and 13 cancer in pleomorphic adenomas. The correlation between CD44v6 and COX-2 and the recurrence and deterioration of pleomorphic adenomas was evaluated using SPSS 15.0 software package. RESULTS: The positive expression rate of CD44v6 in normal parotid gland, primary pleomorphic adenoma, first recurrent pleomorphic adenoma, second recurrent pleomorphic adenoma and cancer was 5.26%, 44.00%, 60.00%, 93.37% and 100.00%, respectively;and the positive expression rate of COX-2 was 10.53%, 40.00%, 63.33%, 93.37% and 100.00%, respectively. There was significant difference in the positive expression intensity of CD44v6 and COX-2 among the groups (P<0.05). The positive expression rate and intensity of CD44v6 and COX-2 in normal parotid gland tissue were significantly lower than those in other groups (P<0.05). There was no significant difference in the positive expression rate and intensity of CD44v6 between primary pleomorphic adenoma and first recurrent pleomorphic adenoma (P>0.05). There was no significant difference in the expression intensity of COX-2 between the two groups (P>0.05), but the positive expression rate of COX-2 between the two groups was not significant(P>0.05). The positive expression rate and intensity of CD44v6 and COX-2 in the second recurrent pleomorphic adenoma were significantly higher than those in the first recurrent pleomorphic adenoma(P<0.05), but there was no significant difference between the two groups(P>0.05). CD44v6 in the second recurrent pleomorphic adenoma and cancer was significantly higher than that in the first recurrent pleomorphic adenoma(P<0.05). There was no significant difference in the expression rate and intensity of COX-2 (P>0.05). CONCLUSIONS: CD44v6 and COX-2 may play an important role in promoting occurrence, invasion, recurrence and deterioration of pleomorphic adenoma through synergistic action.

Poly(norepinephrine)-coated FeOOH nanoparticles as carriers of artemisinin for cancer photothermal-chemical combination therapy.

The photothermal-chemical combination therapy is a promising approach for cancer treatment, however, chemotherapy often causes severe toxic and side effects on normal tissues. Herein, tumor-specific FeOOH@PNE-Art nanoparticles were fabricated via coating poly(norepinephrine) (PNE) on FeOOH nanoparticles, followed by loading of artemisinin (Art). The as-prepared nanoparticles exhibited excellent biocompatibility, strong near-infrared (NIR) absorbance and pH-responsive synchronous release of Art and iron ions. The released iron ions could not only supply iron ions in cancer cells which mediate endoperoxide bridge cleavage of Art and generate reactive oxygen species (ROS), but also react with H2O2 at tumour sites via the Fenton reaction and produce hydroxyl radicals, inducing a tumour-specific killing. Moreover, owing to the synchronous release of Art and iron ions as well as the low leakage of iron ions, FeOOH@PNE-Art nanoparticles showed extremely low toxicity to normal tissue. Under NIR light irradiation, the tumours in FeOOH@PNE-Art injected mice were thoroughly eliminated after 7 days of treatment and no tumour recurrence was found 30 days after treatment, manifesting very high efficacy of combination therapy.

Investigation of Plant Species with Identified Seed Oil Fatty Acids in Chinese Literature and Analysis of Five Unsurveyed Chinese Endemic Species.

Diverse fatty acid structures from different plant species are important renewable resources for industrial raw materials and as liquid fuels with high energy density. Because of its immense geographical and topographical variations, China is a country with enormous diversity of plant species, including large numbers of plants endemic to China. The richness of this resource of species provides a wide range of fatty acids in seeds or other tissues, many of which have been identified by Chinese scientists. However, in the past, most publications describing analysis of these plants were written in Chinese, making access for researchers from other countries difficult. In this study, we investigated reports on seed and fruit oil fatty acids as described in Chinese literature. Six books and more than one thousand papers were collected and the identified fatty acids and relevant plant species were summarized. In total, about 240 fatty acids from almost 1,500 plant species were identified from available Chinese literature. Only about one third of these species were retrieved in the PhyloFAdb and SOFA online databases of plant fatty acids. By referring to a summary of plant species endemic to China, 277 Chinese endemic species from 68 families have been surveyed for seed fatty acids. These account for <2% of total Angiosperm species endemic to China indicating the scope of species yet to be surveyed. To discover additional new fatty acid structures that might benefit society, it is important in the future to study oilseed fatty acids of the many other Chinese endemic plants. As an example, seeds of five unsurveyed species were collected and their fatty acids were analyzed. Ricinoleic acid was detected for the first time in the Salicaceae family.

Improvement of betulinic acid biosynthesis in yeast employing multiple strategies.

BACKGROUND: Betulinic acid (BA) is a lupane-type triterpene which has been considered as a promising agent to cure melanoma with no side effects. Considering that BA is naturally produced in small quantities in plants, we previously reported the success in engineering its production in yeast. In the present study, we attempted to improve BA biosynthesis in yeast by the use of different strategies. RESULTS: We first isolated a gene encoding a lupeol C-28 oxidase (LO) from Betula platyphylla (designated as BPLO). BPLO showed a higher activity in BA biosynthesis compared to the previously reported LOs. In addition, two yeast platforms were compared for engineering the production of BA, which demonstrated that the WAT11 strain was better to host BA pathway than the CEN.PK strain. Based on the WAT11-chassiss, the Gal80p mutant was further constructed. The mutant produced 0.16 mg/L/OD600 of BA, which was 2.2 fold of that produced by the wild type strain (0.07 mg/L/OD600). CONCLUSIONS: This study reported our efforts to improve BA production in yeast employing multiple strategies, which included the identification of a novel LO enzyme with a higher activity in BA biosynthesis, the evaluation of two yeast strains for hosting the BA pathway, and the up-regulation of the expression of the BA pathway genes by managing yeast GAL gene regulon circuit.

[Protective effect of hydrogen sulfide on rats with myocardial ischemia/reperfusion injury and its mechanism].

OBJECTIVE: To explore the protective effect of hydrogen sulfide (H2S) on rats with myocardial ischemia/reperfusion injury (MIRI) and its related mechanism. METHODS: Thirty male SD rats were randomly divided into 3 groups: control group, model group and H2S-treated group (n=10 in each). The MIRI model was established by ligating left anterior descending coronary artery for 40 minutes followed by reperfusion for 2 hours. Rats in H2S-treated group were injected with 14 µmol/kg sodium hydrosulfide (NaHS) intraperitoneally before ischemia and reperfusion, respectively. After 24 hours of reperfusion, blood was collected from abdominal aorta and serum was separated. Then, the heart was removed. The myocardial histopathological changes were observed using HE staining. Serum levels of creatine kinase MB isozyme (CK-MB), lactate dehydrogenase (LDH) and cardiac troponin I (cTnI), as well as the activities of malondialdehyde (MDA), superoxide dismutase (SOD), glutathione-peroxidase (GSH-Px) and catalase (CAT) in the homogenate of myocardium were examined using the kits. Myocardial interleukin (IL)-1ß, IL-6, IL-18 and tumor necrosis factor-α (TNF-α) contents were detected by ELISA. The levels of intracytoplasm Keap 1 and intranuclear nuclear factor-erythroid 2-related factor 2 (Nrf2) and nuclear factor-κB (NF-κB) p65 in myocardium were measured using Western blotting. RESULTS: H2S administration markedly relieved the pathological injury of myocardium. In comparison with control group, serum CK-MB, LDH and cTnI levels, myocardial MDA activities, nuclear factor erythroid 2-related factor 2 (Nrf2) and NF-κB p65 levels in the nucleus, and myocardial contents of IL-1ß, IL-6, IL-18 and TNF-α were significantly elevated, whereas myocardial SOD, GSH-Px and CAT activities as well as Keap 1 level in the cytoplasm were significantly reduced in model group. After H2S treatment, serum CK-MB, LDH and cTnI levels, myocardial MDA activities, Keap 1 level in the cytoplasm, myocardial contents of IL-1ß , IL-6, IL-18 and TNF-α, and NF-κB p65 levels in the nucleus decreased, but myocardial SOD, GSH-Px and CAT activities, and Keap 1 level in the cytoplasm increased as compared with model group. CONCLUSION: Exogenous H2S could confer protection against MIRI in mice. The mechanism might be related to the enhancement of antioxidative ability and the decrease of inflammatory factor release.

Modulating betulinic acid production in Saccharomyces cerevisiae by managing the intracellular supplies of the co-factor NADPH and oxygen.

Betulinic acid is a plant derived triterpenoid with beneficial effects for anti-tumor and anti-human immunodeficiency virus treatments. In Saccharomyces cerevisiae, we previously constructed the betulinic acid biosynthetic pathway, in which several enzymes function in a NADPH or oxygen-dependent manner. To seek whether the intracellular supply of the NADPH/oxygen of S. cerevisiae could be managed for improving betulinic acid production, the expressions of the mutated 2,3-butanediol dehydrogenase (mBDH1) and the yeast codon optimized Vitreoscilla hemoglobin (mvhb) were separately introduced into the betulinic acid forming yeast strain. The effect of these expressions on betulinic acid productivity was evaluated. Our results showed that the expression of mBDH1 and mvhb increased the concentration of betulinic acid to 1.5 and 3.2 times, respectively relative to the controls. Meanwhile, the growth property of these engineered yeast strains was also monitored. Though the mvhb expression greatly improved the production of betulinic acid but exerted a serious inhibition on yeast growth. However, it was possible to keep desirable yeast growth phenotype using an appropriate concentration of acetoin with the expression of mBDH1. The results of this study would provide a general reference to modulate the production of other triterpenoids in S. cerevisiae by managing the supplies of NADPH and oxygen.

The Cysteine2/Histidine2-Type Transcription Factor ZINC FINGER OF ARABIDOPSIS THALIANA6 Modulates Biotic and Abiotic Stress Responses by Activating Salicylic Acid-Related Genes and C-REPEAT-BINDING FACTOR Genes in Arabidopsis.

The cysteine2/histidine2-type zinc finger proteins are a large family of transcription regulators, and some of them play essential roles in plant responses to biotic and abiotic stress. In this study, we found that expression of C2H2-type ZINC FINGER OF ARABIDOPSIS THALIANA6 (AtZAT6) was transcriptionally induced by salt, dehydration, cold stress treatments, and pathogen infection, and AtZAT6 was predominantly located in the nucleus. AtZAT6-overexpressing plants exhibited improved resistance to pathogen infection, salt, drought, and freezing stresses, while AtZAT6 knockdown plants showed decreased stress resistance. AtZAT6 positively modulates expression levels of stress-related genes by directly binding to the TACAAT motifs in the promoter region of pathogen-related genes (ENHANCED DISEASE SUSCEPTIBILITY1, PHYTOALEXIN DEFICIENT4, PATHOGENESIS-RELATED GENE1 [PR1], PR2, and PR5) and abiotic stress-responsive genes (C-REPEAT-BINDING FACTOR1 [CBF1], CBF2, and CBF3). Moreover, overexpression of AtZAT6 exhibited pleiotrophic phenotypes with curly leaves and small-sized plant at vegetative stage and reduced size of floral organs and siliques at the reproductive stage. Modulation of AtZAT6 also positively regulates the accumulation of salicylic acid and reactive oxygen species (hydrogen peroxide and superoxide radical). Taken together, our findings indicated that AtZAT6 plays important roles in plant development and positively modulates biotic and abiotic stress resistance by activating the expression levels of salicylic acid-related genes and CBF genes.

Increase of betulinic acid production in Saccharomyces cerevisiae by balancing fatty acids and betulinic acid forming pathways.

Betulinic acid is a plant-based triterpenoid that has been recognized for its antitumor and anti-HIV activities. The level of betulinic acid in its natural hosts is extremely low. In the present study, we constructed betulinic acid biosynthetic pathway in Saccharomyces cerevisiae by metabolic engineering. Given the betulinic acid forming pathways sharing the common substrate acetyl-CoA with fatty acid synthesis, the metabolic fluxes between the two pathways were varied by changing gene expressions, and their effects on betulinic acid production were investigated. We constructed nine S. cerevisiae strains representing nine combinations of the flux distributions between betulinic acid and fatty acid pathways. Our results demonstrated that it was possible to improve the betulinic acid production in S. cerevisiae while keeping a desirable growth phenotype by optimally balancing the carbon fluxes of the two pathways. Through modulating the expressions of the key genes on betulinic acid and fatty acid pathways, the difference in betulinic acid yield varied largely in the range of 0.01-1.92 mg L(-1) OD(-1). The metabolic engineering approach used in this study could be extended for synthesizing other triterpenoids in S. cerevisiae.

Identifying three ecological chemotypes of Xanthium strumarium glandular trichomes using a combined NMR and LC-MS method.

Xanthanolides, as the sesquiterpene lactones, are reportedly the major components for the pharmacological properties of X. strumarium L. species. Phytochemical studies indicated that the glandular structures on the surface of plant tissues would form the primary sites for the accumulation of this class of the compounds. As the interface between plants and their natural enemies, glandular trichomes may vary with respect to which of their chemicals are sequestered against different herbivores in different ecologies. However, to date, no data are available on the chemical characterisation of X. strumarium glandular cells. In this study, the trichome secretions of the X. strumarium species originating from nineteen unique areas across eleven provinces in China, were analysed by HPLC, LC-ESI-MS and NMR. For the first time three distinct chemotypes of X. strumarium glandular trichomes were discovered along with the qualitative and quantitative evaluations of their presence of xanthanolides; these were designated glandular cell Types I, II, and III, respectively. The main xanthanolides in Type I cells were 8-epi-xanthatin and xanthumin while no xanthatin was detected. Xanthatin, 8-epi-xanthatin, and xanthumin dominated in Type II cells with comparable levels of each being present. For Type III cells, significantly higher concentrations of 8-epi-xanthatin or xanthinosin (relative to xanthatin) were detected with xanthinosin only being observed in this type. Further research will focus on understanding the ecological and molecular mechanism causing these chemotype differences in X. strumarium glandular structures.

Molecular characterization of the pentacyclic triterpenoid biosynthetic pathway in Catharanthus roseus.

Catharanthus roseus is an important medicinal plant and the sole commercial source of monoterpenoid indole alkaloids (MIA), anticancer compounds. Recently, triterpenoids like ursolic acid and oleanolic acid have also been found in considerable amounts in C. roseus leaf cuticular wax layer. These simple pentacyclic triterpenoids exhibit various pharmacological activities such as anti-inflammatory, anti-tumor and anti-microbial properties. Using the EST collection from C. roseus leaf epidermome ( http://www.ncbi.nlm.nih.gov/dbEST ), we have successfully isolated a cDNA (CrAS) encoding 2,3-oxidosqualene cyclase (OSC) and a cDNA (CrAO) encoding amyrin C-28 oxidase from the leaves of C. roseus. The functions of CrAS and CrAO were analyzed in yeast (Saccharomyces cerevisiae) systems. CrAS was characterized as a novel multifunctional OSC producing α- and ß-amyrin in a ratio of 2.5:1, whereas CrAO was a multifunctional C-28 oxidase converting α-amyrin, ß-amyrin and lupeol to ursolic-, oleanolic- and betulinic acids, respectively, via a successive oxidation at the C-28 position of the substrates. In yeast co-expressing CrAO and CrAS, ursolic- and oleanolic acids were detected in the yeast cell extracts, while the yeast cells co-expressing CrAO and AtLUP1 from Arabidopsis thaliana produced betulinic acid. Both CrAS and CrAO genes show a high expression level in the leaf, which was consistent with the accumulation patterns of ursolic- and oleanolic acids in C. roseus. These results suggest that CrAS and CrAO are involved in the pentacyclic triterpene biosynthesis in C. roseus.