Metabolic engineering Corynebacterium glutamicum for D-chiro-inositol production.

D-chiro-inositol (DCI) is a potential drug for the treatment of type II diabetes and polycystic ovary syndrome. In order to effectively synthesize DCI in Corynebacterium glutamicum, the genes related to inositol catabolism in clusters iol1 and iol2 were knocked out in C. glutamicum SN01 to generate the chassis strain DCI-1. DCI-1 did not grow in and catabolize myo-inositol (MI). Subsequently, different exogenous and endogenous inosose isomerases were expressed in DCI-1 and their conversion ability of DCI from MI were compared. After fermentation, the strain DCI-7 co-expressing inosose isomerase IolI2 and inositol dehydrogenase IolG was identified as the optimal strain. Its DCI titer reached 3.21 g/L in the presence of 20 g/L MI. On this basis, the pH, temperature and MI concentration during whole-cell conversion of DCI by strain DCI-7 were optimized. Finally, the optimal condition that achieved the highest DCI titer of 6.96 g/L were obtained at pH 8.0, 37 °C and addition of 40 g/L MI. To our knowledge, it is the highest DCI titer ever reported.

Toxicological, biochemical, and in silico investigations of three trehalase inhibitors for new ways to control aphids.

Insect trehalases have been identified as promising new targets for pest control. These key enzymes are involved in trehalose hydrolysis and plays an important role in insect growth and development. In this contribution, plant and microbial compounds, namely validamycin A, amygdalin, and phloridzin, were evaluated for their effect, through trehalase inhibition, on Acyrthosiphon pisum aphid. The latter is part of the Aphididae family, main pests as phytovirus vectors and being very harmful for crops. Validamycin A was confirmed as an excellent trehalase inhibitor with an half maximal inhibitory concentration and inhibitor constant of 2.2 × 10-7 and 5 × 10-8 M, respectively, with a mortality rate of ~80% on a A. pisum population. Unlike validamycin A, the insect lethal efficacy of amygdalin and phloridzin did not correspond to their trehalase inhibition, probably due to their hydrolysis by insect ß-glucosidases. Our docking studies showed that none of the three compounds can bind to the trehalase active site, unlike their hydrolyzed counterparts, that is, validoxylamine A, phloretin, and prunasin. Validoxylamine A would be by far the best trehalase binder, followed by phloretin and prunasin.

D-Pinitol mitigates post-traumatic stress disorder-like behaviors induced by single prolonged stress in mice through mineralocorticoid receptor antagonism.

Post-traumatic stress disorder (PTSD) is a mental illness that can occur in individuals who have experienced trauma. Current treatments for PTSD, typically serotonin reuptake inhibitors, have limited effectiveness for patients and often cause serious adverse effects. Therefore, a novel class of treatment with better pharmacological profile is necessary. D-Pinitol has been reported to be effective for depression and anxiety disorders, but there are no reports associated with PTSD. In the present study, we investigated the effects of D-pinitol in a mouse model of PTSD induced by a single prolonged stress (SPS) protocol. We examined the therapeutic effects of D-pinitol on emotional and cognitive impairments in the SPS mouse model. We also investigated the effects of D-pinitol on fear memory formation. Mineralocorticoid receptor transactivation assay, Western blot, and quantitative PCR were employed to investigate how D-pinitol exerts its pharmacological activities. D-Pinitol ameliorated PTSD-like behaviors in a SPS mouse model. D-Pinitol also normalized the increased mRNA expression levels and protein levels of the mineralocorticoid receptor in the amygdala. A mineralocorticoid receptor agonist reversed the effects of D-pinitol on fear extinction and recall, and the antagonistic property of D-pinitol against the mineralocorticoid receptor was confirmed in vitro. Our findings suggest that D-pinitol could serve as a potential therapeutic agent for PTSD due to its antagonistic effect on the mineralocorticoid receptor.

Changes in primary metabolite content may affect thrips feeding preference in soybean crops.

Past research has characterized the induction of plant defenses in response to chewing insect damage. However, little is known about plant responses to piercing-sucking insects that feed on plant cell-contents like thrips (Caliothrips phaseoli). In this study, we used NMR spectroscopy to measure metabolite changes in response to six days of thrips damage from two field-grown soybean cultivars (cv.), known for their different susceptibility to Caliothrips phaseoli. We observed that thrips damage reduces sucrose concentration in both cultivars, while pinitol, the most abundant leaf soluble carbohydrate, is induced in cv. Charata but not in cv. Williams. Thrips did not show preference for leaves where sucrose or pinitol were externally added, at tested concentration. In addition, we also noted that cv. Charata was less naturally colonized and contained higher levels of trigonelline, tyrosine as well as several compounds that we have not yet identified. We have established that preference-feeding clues are not dependent on the plants major soluble carbohydrates but may depend on other types of compounds or leaf physical characteristics.

D-pinitol alleviates diabetic cardiomyopathy by inhibiting the optineurin-mediated endoplasmic reticulum stress and glycophagy signaling pathway.

Diabetic cardiomyopathy (DCM) is an important complication resulting in heart failure and death of diabetic patients. However, there is no effective drug for treatments. This study investigated the effect of D-pinitol (DP) on cardiac injury using diabetic mice and glycosylation injury of cardiomyocytes and its molecular mechanisms. We established the streptozotocin-induced SAMR1 and SAMP8 mice and DP (150 mg/kg/day) intragastrically and advanced glycation end-products (AGEs)-induced H9C2 cells. H9C2 cells were transfected with optineurin (OPTN) siRNA and overexpression plasmids. The metabolic disorder indices, cardiac dysfunction, histopathology, immunofluorescence, western blot, and immunoprecipitation were investigated. Our results showed that DP reduced the blood glucose and AGEs, and increased the expression of heart OPTN in diabetic mice and H9C2 cells, thereby inhibiting the endoplasmic reticulum stress (GRP78, CHOP) and glycophagy (STBD1, GABARAPL1), and alleviating the myocardial apoptosis and fibrosis of DCM. The expression of filamin A as an interaction protein of OPTN downregulated by AGEs decreased OPTN abundance. Moreover, OPTN siRNA increased the expression of GRP78, CHOP, STBD1, and GABARAPL1 and inhibited the expression of GAA via GSK3ß phosphorylation and FoxO1. DP may be helpful to treat the onset of DCM. Targeting OPTN with DP could be translated into clinical application in the fighting against DCM.

In vivo evaluation of the neurogenotoxic effects of exposure to validamycin A in neuroblasts of Drosophila melanogaster larval brain.

Agriculture commonly utilizes crop protection products to tackle infestations from fungi, parasites, insects, and weeds. Validamycin A, an inhibitor of trehalase, possesses antibiotic and antifungal attributes. Epidemiological evidence has led to concerns regarding a potential link between pesticide usage and neurodegenerative diseases. The fruit fly, Drosophila melanogaster, has been recognized as a reliable model for genetic research due to its significant genetic similarities with mammals. Here, we propose to use D. melanogaster as an effective in vivo model system to investigate the genotoxic risks associated with exposure to validamycin A. In this study, we performed a neurotoxic evaluation of validamycin A in D. melanogaster larvae. Several endpoints were evaluated, including toxicity, intracellular oxidative stress (reactive oxygen species), intestinal damage, larval behavior (crawling behavior, light/dark sensitivity assay, and temperature sensitivity assay), locomotor (climbing) behavior, and neurogenotoxic effects (impaired DNA via Comet assay, enhanced by Endo III and formamidopyrimidine DNA glycosylase [FPG]). The results showed that exposure to validamycin A, especially at higher doses (1 and 2.5 mM), induced DNA impairment in neuroblasts as observed by Comet assay. Both larvae and adults exhibited behavioral changes and produced reactive oxygen species. Most importantly, this research represents a pioneering effort to report neurogenotoxicity data specifically in Drosophila larval neuroblasts, thus underscoring the importance of this species as a testing model in exploring the biological impacts of validamycin A. The in vivo findings from the experiments are a valuable and novel addition to the existing validamycin A neurogenotoxicity database.

Comparative effects of fixed-dose mitiglinide/voglibose combination and glimepiride on vascular endothelial function and glycemic variability in patients with type 2 diabetes: A randomized controlled trial.

INTRODUCTION: The aim of this study was to compare the effects of mitiglinide/voglibose with those of glimepiride on glycemic variability and vascular endothelial function in patients with type 2 diabetes. MATERIALS AND METHODS: It was a multicenter, open-label, randomized, crossover study. Hospitalized patients received either mitiglinide/voglibose (three times daily administration of 10 mg mitiglinide and 0.2 mg voglibose) or glimepiride (once-daily 2 mg) in random order, each for 5 days. The reactive hyperemia index (RHI) and the mean amplitude of glycemic excursions (MAGE) were measured as co-primary endpoints using reactive hyperemia peripheral arterial tonometry and continuous glucose monitoring. RESULTS: The analysis included 30 patients (15 in each group). The RHI was 1.670 ± 0.369 during treatment with mitiglinide/voglibose and 1.716 ± 0.492 during treatment with glimepiride, with no significant difference between the two. MAGE was significantly lower in the mitiglinide/voglibose group (47.6 ± 18.5 mg/dL) than in the glimepiride group (100.6 ± 32.2 mg/dL). Although the mean blood glucose levels over the entire 24 h period were comparable between the two groups, the use of mitiglinide/voglibose was associated with a lower standard deviation of mean glucose, coefficient of variation, and mean postprandial glucose excursion compared with glimepiride. The time below range (<70 mg/dL) and the time above range (>180, >200, and 250 mg/dL) were lower in the mitiglinide/voglibose group, while the time in range (70-180 mg/dL) was higher. CONCLUSIONS: In our short-duration randomized crossover study, although not impacting vascular endothelial function, mitiglinide/voglibose demonstrated potential benefits in reducing glycemic variability, postprandial hyperglycemia, and hypoglycemia in patients with type 2 diabetes.

Pharmacokinetics and Endocrine Effects of an Oral Dose of D-Pinitol in Human Fasting Healthy Volunteers.

The present study characterizes the oral pharmacokinetics of D-Pinitol, a natural insulin mimetic inositol, in human healthy volunteers (14 males and 11 females). D-Pinitol absorption was studied in (a) subjects receiving a single oral dose of 15 mg/kg (n = 10), or (b) 5 mg/kg pure D-Pinitol (n = 6), and (c) subjects receiving D-Pinitol as part of carbohydrate-containing carob pods-derived syrup with a 3.2% D-Pinitol (Dose of 1600 mg/subject, n = 9). The volunteers received a randomly assigned single dose of either D-Pinitol or carob pod-derived syrup. Blood samples were collected at 0, 15, 30, 45, 60, 90, 120, 180, 240, 360 and 1440 min after intake. Plasma concentration of D-Pinitol was measured and pharmacokinetic parameters obtained. The data indicate that when given alone, the oral absorption of D-Pinitol is dose-dependent and of extended duration, with a Tmax reached after almost 4 h, and a half-life greater than 5 h. When the source of D-Pinitol was a carob pods-derived syrup, Cmax was reduced to 40% of the expected based on the data of D-Pinitol alone, suggesting a reduced absorption probably because of competition with monosaccharide transport. In this group, Tmax was reached before that of D-Pinitol alone, but the estimated half-life remained the same. In the D-Pinitol groups, plasma concentrations of glucose, insulin, glucagon, ghrelin, free fatty acids, and pituitary hormones were additionally measured. A dose of 15 mg/kg of D-Pinitol did not affect glucose levels in healthy volunteers, but reduced insulin and increased glucagon and ghrelin concentrations. D-Pinitol did not increase other hormones known to enhance plasma glucose, such as cortisol or GH, which were surprisingly reduced after the ingestion of this inositol. Other pituitary hormones (gonadotropins, prolactin, and thyroid-stimulating hormone) were not affected after D-Pinitol ingestion. In a conclusion, D-Pinitol is absorbed through the oral route, having an extended half-life and displaying the pharmacological profile of an endocrine pancreas protector, a pharmacological activity of potential interest for the treatment or prevention of insulin resistance-associated conditions.

Effects of luseogliflozin and voglibose on high-risk lipid profiles and inflammatory markers in diabetes patients with heart failure.

Sodium-glucose cotransporter 2 inhibitors could reduce cardiovascular events in patients with heart failure irrespective of diabetes status. In this prespecified sub-analysis of randomised-controlled trial, we investigated the efficacy of luseogliflozin (2.5 mg daily), a sodium-glucose cotransporter 2 inhibitor, with that of voglibose (0.6 mg daily), an alpha-glucosidase inhibitor, on high-risk lipid profile and inflammatory markers in patients with type-2 diabetes and heart failure. Among the 157 patients studied, there were no significant differences in the mean malondialdehyde LDL or small-dense LDL cholesterol levels between the luseogliflozin and voglibose groups (percent change: 0.2% vs. - 0.6%, p = 0.93; - 1.7% vs. - 8.6%, p = 0.21) after 12 weeks in comparison to levels at the baseline. No significant difference was observed between the two groups in the adiponectin and high-sensitivity C-reactive protein levels after 12 weeks compared to the baseline levels (percent change, - 1.6% vs. - 4.0% and 22.5% vs. 10.0%; p = 0.52 and p = 0.55, respectively). In conclusion, in patients with type-2 diabetes and heart failure, compared to voglibose, luseogliflozin did not significantly improve the high-risk lipoprotein profile including malondialdehyde LDL and small-dense LDL cholesterol or the levels of inflammatory markers, including adiponectin and high-sensitivity C-reactive protein.Trial registration: Trial number: UMIN-CTR, UMIN000018395; Registered 23 July 2015; URL: https://www.umin.ac.jp/ctr/index.htm .

The effects of the voglibose on non-alcoholic fatty liver disease in mice model.

The α-glucosidase inhibitor (α-GI) delays the intestinal absorption of glucose, which reduces postprandial hepatic glucose intake. This mechanism is considered to be effective in treating non-alcoholic fatty liver disease (NAFLD). Here, we investigated the effect of voglibose, an α-glucosidase inhibitor, on high-fat, high-fructose (HFHFr) diet-induced NAFLD models. Seven-week-old male C57BL/6J mice were randomly placed in a chow diet group or an HFHFr diet group. After 10 weeks, mice in the HFHFr group were randomly assigned to one of three groups: HFHFr diet with vehicle, HFHFr with voglibose, or HFHFr with pioglitazone. Each diet and treatment was continued for 10 weeks. The HFHFr diet induced severe NAFLD in terms of steatosis, hepatitis, and fibrosis. Administration of voglibose improved all aspects of NAFLD, comparable to those of pioglitazone, a positive control. In voglibose-treated mice, gene expressions of hepatic lipogenesis markers were significantly downregulated. In the in vitro experiment, reducing the influx of glucose into hepatocytes significantly reduced steatosis and de novo lipogenesis even in the presence of sufficient fructose and fat, demonstrating that the mechanism of voglibose could be effective in treating HFHFr diet-induced NAFLD. These results indicate that voglibose improves HFHFr diet-induced NAFLD by suppressing hepatic de novo lipogenesis.

Hazard Assessment of the Effects of Acute and Chronic Exposure to Permethrin, Copper Hydroxide, Acephate, and Validamycin Nanopesticides on the Physiology of Drosophila: Novel Insights into the Cellular Internalization and Biological Effects.

New insights into the interactions between nanopesticides and edible plants are required in order to elucidate their impacts on human health and agriculture. Nanopesticides include formulations consisting of organic/inorganic nanoparticles. Drosophila melanogaster has become a powerful model in genetic research thanks to its genetic similarity to mammals. This project mainly aimed to generate new evidence for the toxic/genotoxic properties of different nanopesticides (a nanoemulsion (permethrin nanopesticides, 20 ± 5 nm), an inorganic nanoparticle as an active ingredient (copper(II) hydroxide [Cu(OH)2] nanopesticides, 15 ± 6 nm), a polymer-based nanopesticide (acephate nanopesticides, 55 ± 25 nm), and an inorganic nanoparticle associated with an organic active ingredient (validamycin nanopesticides, 1177 ± 220 nm)) and their microparticulate forms (i.e., permethrin, copper(II) sulfate pentahydrate (CuSO4·5H2O), acephate, and validamycin) widely used against agricultural pests, while also showing the merits of using Drosophila-a non-target in vivo eukaryotic model organism-in nanogenotoxicology studies. Significant biological effects were noted at the highest doses of permethrin (0.06 and 0.1 mM), permethrin nanopesticides (1 and 2.5 mM), CuSO4·5H2O (1 and 5 mM), acephate and acephate nanopesticides (1 and 5 mM, respectively), and validamycin and validamycin nanopesticides (1 and 2.5 mM, respectively). The results demonstrating the toxic/genotoxic potential of these nanopesticides through their impact on cellular internalization and gene expression represent significant contributions to future nanogenotoxicology studies.

D-pinitol attenuates isoproterenol-induced myocardial infarction by alleviating cardiac inflammation, oxidative stress and ultrastructural changes in Swiss albino mice.

Cardiovascular diseases are the most disturbing problems throughout the world. The side effects of existing drugs are continuously compelling the scientist to look for better options in terms of safety, efficacy and cost-effectiveness. Our study is also a move in this direction. We have chosen D-pinitol to see its cardioprotective role in isoproterenol-induced myocardial infarction in Swiss albino mice. Grouping was made by dividing mice into eight groups (n = 6). Group I, control; Group II, isoproterenol (ISO) (150 mg/kg, i.p.); Group III, D-pinitol (PIN) (25 mg); Group IV, PIN (50 mg); Group V, PIN (100 mg) per kg per oral, respectively with ISO; Group VI, PIN per se (100 mg D-pinitol only); Group VII, Propranolol (PRO) (20 mg/kg/oral) with ISO; and Group VIII, PRO per se (20 mg/kg, p.o.). After 24 h of the last dose, the blood sample was collected for biochemical parameters, then mice were, killed through cervical dislocation under anaesthesia and cardiac tissue was collected for biochemical, histopathological and ultrastructural evaluation. Administration of ISO in mice altered the level of antioxidant markers, cardiac injury markers and inflammatory markers, which were significantly restored towards normal by D-pinitol at the dose of 50 and 100 mg. 25 mg of D-pinitol dosage, did not produce significant cardio protection. The histopathological and ultrastructural analysis further confirmed these findings. Our study showed that D-pinitol significantly protected myocardial damage which was induced by ISO and reverted oxidative stress and inflammation considerably.

Seventy-Two-Hour LRRK2 Kinase Activity Inhibition Increases Lysosomal GBA Expression in H4, a Human Neuroglioma Cell Line.

Mutations in LRRK2 and GBA1 are key contributors to genetic risk of developing Parkinson's disease (PD). To investigate how LRRK2 kinase activity interacts with GBA and contributes to lysosomal dysfunctions associated with the pathology of PD. The activity of the lysosomal enzyme ß-Glucocerebrosidase (GCase) was assessed in a human neuroglioma cell model treated with two selective inhibitors of LRKK2 kinase activity (LRRK2-in-1 and MLi-2) and a GCase irreversible inhibitor, condutirol-beta-epoxide (CBE), under 24 and 72 h experimental conditions. We observed levels of GCase activity comparable to controls in response to 24 and 72 h treatments with LRRK2-in-1 and MLi-2. However, GBA protein levels increased upon 72 h treatment with LRRK2-in-1. Moreover, LC3-II protein levels were increased after both 24 and 72 h treatments with LRRK2-in-1, suggesting an activation of the autophagic pathway. These results highlight a possible regulation of lysosomal function through the LRRK2 kinase domain and suggest an interplay between LRRK2 kinase activity and GBA. Although further investigations are needed, the enhancement of GCase activity might restore the defective protein metabolism seen in PD.

Pharmacological effects of D-Pinitol - A comprehensive review.

In recent years, the application of phytochemicals to prevent or treat diseases has received greater attention. These phytochemicals have little or no toxicity against healthy tissues and are thus considered as ideal compounds. An impressive number of modern drugs are obtained from natural sources based on their traditional value. D-Pinitol is a natural compound that is derived from soy and soy products. It is a potentially active molecule that belongs to the class of inositols. D-pinitol has been pharmacologically evaluated for its potent antioxidant, anti-diabetic, anti-inflammatory, anti-cancer, hepatoprotective, cardioprotective, renoprotective, neuroprotective, immunosuppressive, and anti-osteoporotic efficacies. This review is an attempt to validate the plausible pharmacological effects of D-pinitol using various in vivo and in vitro studies. PRACTICAL IMPLICATIONS: The consumption of plant-based products has been significantly increased all over the world. The active phytochemicals that are found in plants are stated to have numerous health promoting functions for the treatment of diabetes, cancer, inflammation, cardiac diseases, liver dysfunction, and many other. D-Pinitol is abundantly present in soybeans that possess notable therapeutic activities. Understanding the effects of D-Pinitol would potentially help in applying this compound in clinical research for the treatment of different disorders.

d-Pinitol promotes tau dephosphorylation through a cyclin-dependent kinase 5 regulation mechanism: A new potential approach for tauopathies?

BACKGROUND AND PURPOSE: Recent evidence links brain insulin resistance with neurodegenerative diseases, where hyperphosphorylated tau protein contributes to neuronal cell death. In the present study, we aimed to evaluate if d-pinitol inositol, which acts as an insulin sensitizer, affects the phosphorylation status of tau protein. EXPERIMENTAL APPROACH: We studied the pharmacological effect of d-pinitol on insulin signalling and tau phosphorylation in the hippocampus of Wistar and Zucker rats. To this end, we evaluated by western blotting the Akt pathway and its downstream proteins as being one of the main insulin-mediator pathways. Also, we explored the functional status of additional kinases phosphorylating tau, including PKA, ERK1/2, AMPK and CDK5. We utilized the 3xTg mouse model as a control for tauopathy, since it carries tau mutations that promote phosphorylation and aggregation. KEY RESULTS: Surprisingly, we discovered that oral d-pinitol treatment lowered tau phosphorylation significantly, but not through the expected kinase GSK-3 regulation. An extensive search for additional kinases phosphorylating tau revealed that this effect was mediated through a mechanism dependent on the reduction of the activity of the CDK5, affecting both its p35 and p25 subunits. This effect disappeared in leptin-deficient Zucker rats, uncovering that the association of leptin deficiency, obesity, dyslipidaemia and hyperinsulinaemia abrogates d-pinitol actions on tau phosphorylation. The 3xTg mice confirmed d-pinitol effectiveness in a genetic AD-tauopathy. CONCLUSION AND IMPLICATIONS: The present findings suggest that d-pinitol, by regulating CDK5 activity through a decrease of CDK5R1, is a potential drug for developing treatments for neurological disorders such as tauopathies.

d-Chiro-Inositol extends the lifespan of male Drosophila melanogaster better than d-Pinitol through insulin signaling and autophagy pathways.

d-Pinitol (DP) is the methylated product of d-Chiro-Inositol (DCI), which is one of the nine isomers of inositol with optical activity. Both substances possess antioxidant activity. This study was conducted to investigate and compare the antioxidant and life-prolonging effects of DCI and DP on male Drosophila melanogaster. Results showed that DCI and DP prolonged the lifespan and improved the climbing, anti-stress, and antioxidant activities. After treatment with DCI and DP, intestinal homeostasis was improved and the abnormal proliferation of intestinal stem cells (ISCs) was attenuated. Furthermore, real-time PCR revealed downregulated expression levels of PI3K and Akt and upregulated expression levels of Dilp5 and FOXO, which consequently activated Atg1, Atg5, Atg8a, and Atg8b and increased the number of lysosomes. Altogether, DCI exerts a slightly better effect than DP based on various indicators. RNAi D. melanogaster lifespan and molecular docking results further suggested that DCI and DP could prolong longevity through insulin signaling (IIS) and autophagy pathways.

D-Pinitol-Active Natural Product from Carob with Notable Insulin Regulation.

Carob is one of the major food trees for peoples of the Mediterranean basin, but it has also been traditionally used for medicinal purposes. Carob contains many nutrients and active natural products, and D-Pinitol is clearly one of the most important of these. D-Pinitol has been reported in dozens of scientific publications and its very diverse medicinal properties are still being studied. Presently, more than thirty medicinal activities of D-Pinitol have been reported. Among these, many publications have reported the strong activities of D-Pinitol as a natural antidiabetic and insulin regulator, but also as an active anti-Alzheimer, anticancer, antioxidant, and anti-inflammatory, and is also immune- and hepato-protective. In this review, we will present a brief introduction of the nutritional and medicinal importance of Carob, both traditionally and as found by modern research. In the introduction, we will present Carob's major active natural products. The structures of inositols will be presented with a brief literature summary of their medicinal activities, with special attention to those inositols in Carob, as well as D-Pinitol's chemical structure and its medicinal and other properties. D-Pinitol antidiabetic and insulin regulation activities will be extensively presented, including its proposed mechanism of action. Finally, a discussion followed by the conclusions and future vision will summarize this article.

Engineered plant control of associative nitrogen fixation.

Engineering N2-fixing symbioses between cereals and diazotrophic bacteria represents a promising strategy to sustainably deliver biologically fixed nitrogen (N) in agriculture. We previously developed novel transkingdom signaling between plants and bacteria, through plant production of the bacterial signal rhizopine, allowing control of bacterial gene expression in association with the plant. Here, we have developed both a homozygous rhizopine producing (RhiP) barley line and a hybrid rhizopine uptake system that conveys upon our model bacterium Azorhizobium caulinodans ORS571 (Ac) 103-fold improved sensitivity for rhizopine perception. Using this improved genetic circuitry, we established tight rhizopine-dependent transcriptional control of the nitrogenase master regulator nifA and the N metabolism σ-factor rpoN, which drove nitrogenase expression and activity in vitro and in situ by bacteria colonizing RhiP barley roots. Although in situ nitrogenase activity was suboptimally effective relative to the wild-type strain, activation was specific to RhiP barley and was not observed on the roots of wild-type plants. This work represents a key milestone toward the development of a synthetic plant-controlled symbiosis in which the bacteria fix N2 only when in contact with the desired host plant and are prevented from interaction with nontarget plant species.

Spectroscopic and theoretical evidences for the surface binding of voglibose drug with DNA.

Binding of voglibose (VOG), an alpha glucosidase inhibitor, with CT-DNA has been investigated using various spectroscopic techniques including UV-Vis, fluorescence and circular dichroism (CD) coupled with relative viscosity. Isothermal titration calorimetric studies have been used to calculate the thermodynamic parameters such as ΔH (0.0188 cal/mol), ΔS (63.3 cal/mol/K) and ΔG (-18.8 kcal/mol), which reveal that the binding is a spontaneous process and hydrophobic and H-bonding interactions play major roles in the binding process. Effect of ionic strength confirms the existence of hydrophobic interaction between VOG and CT-DNA. Competitive displacement assays with ethidium bromide (EB) and Hoechst 33258 suggest that VOG possibly binds on the surface of CT-DNA. Viscosity measurements also disclose that the binding could be mainly surface binding. Corroborating the experimental observations, metadynamics molecular simulation studies confirm that VOG binding on the surface of the DNA molecule through hydrophobic interactions and direct and water molecule mediated H-bonding.

Pinitol Protects Against Ox-Low-Density Lipoprotein-Induced Endothelial Inflammation and Monocytes Attachment.

ABSTRACT: Atherosclerosis is a cardiovascular disease that affects a majority of people around the world at old age. Atherosclerosis is slow to develop and challenging to treat. Endothelial dysfunction caused by oxidative stress, inflammation, and other pathological factors drives the process of atherogenesis. LOX-1 is one of the main scavenging receptors for oxidized low-density lipoprotein (ox-LDL) and contributes to atherogenesis by inducing overproduction of reactive oxygen species, increased expression of proinflammatory cytokines, and secretion of cellular adhesion molecules. In addition, activation of LOX-1 inhibits the expression of KLF2, a key protective factor against atherosclerosis. In this study, we investigated the effects of pinitol, and naturally occurring cyclic polyol, on endothelial dysfunction induced by ox-LDL. Our findings show that pinitol revealed a good safety profile, as evidenced by reducing lactate dehydrogenase release in human aortic endothelial cells. In our experiments, pinitol reduced the production of reactive oxygen species and expression of IL-6 and monocyte chemoattractant protein-1 induced by ox-LDL. Pinitol also significantly reduced the attachment of THP-1 monocytes to endothelial cells via downregulation of vascular cellular adhesion molecule-1 and E-selectin. Importantly, we found that pinitol reduced the expression of LOX-1 induced by ox-LDL and rescued the expression of KLF2, which is dependent on ERK5 expression. Together, our findings provide notable evidence that pinitol may have potential implication in the prevention and treatment of atherosclerosis.