Greener and sustainable method for alkene epoxidations by polymer-supported Mo(VI) catalysts

A polybenzimidazole supported Mo(VI) (PBI.Mo) catalyst has been prepared and characterised. The catalytic activities of the PBI.Mo catalyst in epoxidation of alkenes with tert-butyl hydroperoxide (TBHP) as an oxidant have been studied under different reaction conditions in a batch reactor. As alkene representatives we have chosen cyclohexene, limonene, α-pinene and 1-octene (a less reactive terminal alkene). The order of reactivity of the alkenes was found to be: cyclohexene>limonene>α-pinene>1-octene. The stability of each polymer catalyst was assessed by recycling a sample in batch reaction using conditions that will form the basis of the continuous process. The loss of Mo from each support has been investigated by isolating any residue from the reaction supernatant solutions, following removal of the heterogeneous polymer catalyst, and then using the residues as potential catalysts in epoxidation reactions

Fungi colonizing the soil and roots of tomato (Lycopersicum esculentum Mill.) plants treated with biological control agents

Tomato plants, cv. Rumba Ożarowska, grown in the greenhouse of the University of Warmia and Mazury, were protected in the form of alternate spraying (twice) and watering (twice) with 5% aqueous extracts of the following plant species: Aloe vulgaris Lam., Achillea millefolium L., Mentha piperita L., Polygonum aviculare L., Equisetum arvense L., Juglans regia L. and Urtica dioica L. Plants not treated with the extracts served as control. After fruit harvest, samples of roots and soil were collected. The roots were disinfected and next placed on PDA medium. Soil-colonizing fungi were cultured on Martin medium. Fungi were identified microscopically after incubation. Pathogenic fungal species, Colletotrichum coccodes, Fusarium equiseti, F. oxysporum and F. poae, accounted for over 60% of all isolates obtained from the roots of tomato plants. The soil fungal community was dominated by yeast-like fungi (75.4%), whereas pathogenic fungi were present in low numbers. The applied 5% aqueous plant extracts effectively reduced the abundance of fungi, including pathogenic species, colonizing tomato plants and soil. The extract from P. aviculare showed the highest efficacy, while the extract from J. regia was least effective. Fungi showing antagonistic activity against pathogens (Paecilomyces roseum and species of the genus Trichoderma) were isolated in greatest abundance from the soil and the roots of tomato plants treated with A. millefolium, M. piperita and U. dioica extracts

Abortive Lytic Reactivation of KSHV in CBF1/CSL Deficient Human B Cell Lines

Since Kaposi's sarcoma associated herpesvirus (KSHV) establishes a persistent infection in human B cells, B cells are a critical compartment for viral pathogenesis. RTA, the replication and transcription activator of KSHV, can either directly bind to DNA or use cellular DNA binding factors including CBF1/CSL as DNA adaptors. In addition, the viral factors LANA1 and vIRF4 are known to bind to CBF1/CSL and modulate RTA activity. To analyze the contribution of CBF1/CSL to reactivation in human B cells, we have successfully infected DG75 and DG75 CBF1/CSL knock-out cell lines with recombinant KSHV.219 and selected for viral maintenance by selective medium. Both lines maintained the virus irrespective of their CBF1/CSL status. Viral reactivation could be initiated in both B cell lines but viral genome replication was attenuated in CBF1/CSL deficient lines, which also failed to produce detectable levels of infectious virus. Induction of immediate early, early and late viral genes was impaired in CBF1/CSL deficient cells at multiple stages of the reactivation process but could be restored to wild-type levels by reintroduction of CBF1/CSL. To identify additional viral RTA target genes, which are directly controlled by CBF1/CSL, we analyzed promoters of a selected subset of viral genes. We show that the induction of the late viral genes ORF29a and ORF65 by RTA is strongly enhanced by CBF1/CSL. Orthologs of ORF29a in other herpesviruses are part of the terminase complex required for viral packaging. ORF65 encodes the small capsid protein essential for capsid shell assembly. Our study demonstrates for the first time that in human B cells viral replication can be initiated in the absence of CBF1/CSL but the reactivation process is severely attenuated at all stages and does not lead to virion production. Thus, CBF1/CSL acts as a global hub which is used by the virus to coordinate the lytic cascade

Age-dependent and sex-dependent disparity in mortality in patients with adrenal incidentalomas and autonomous cortisol secretion: an international, retrospective, cohort study

Background: The association between cortisol secretion and mortality in patients with adrenal incidentalomas is controversial. We aimed to assess all-cause mortality, prevalence of comorbidities, and occurrence of cardiovascular events in uniformly stratified patients with adrenal incidentalomas and cortisol autonomy (defined as non-suppressible serum cortisol on dexamethasone suppression testing). Methods: We conducted an international, retrospective, cohort study (NAPACA Outcome) at 30 centres in 16 countries. Eligible patients were aged 18 years or older with an adrenal incidentaloma (diameter ≥1 cm) detected between Jan 1, 1996, and Dec 31, 2015, and availability of a 1 mg dexamethasone suppression test result from the time of the initial diagnosis. Patients with clinically apparent hormone excess, active malignancy, or follow-up of less than 36 months were excluded. Patients were stratified according to the 0800–0900 h serum cortisol values after an overnight 1 mg dexamethasone suppression test; less than 50 nmol/L was classed as non-functioning adenoma, 50–138 nmol/L as possible autonomous cortisol secretion, and greater than 138 nmol/L as autonomous cortisol secretion. The primary endpoint was all-cause mortality. Secondary endpoints were the prevalence of cardiometabolic comorbidities, cardiovascular events, and cause-specific mortality. The primary and secondary endpoints were assessed in all study participants. Findings: Of 4374 potentially eligible patients, 3656 (2089 [57·1%] with non-functioning adenoma, 1320 [36·1%] with possible autonomous cortisol secretion, and 247 [6·8%] with autonomous cortisol secretion) were included in the study cohort for mortality analysis (2350 [64·3%] women and 1306 [35·7%] men; median age 61 years [IQR 53–68]; median follow-up 7·0 years [IQR 4·7–10·2]). During follow-up, 352 (9·6%) patients died. All-cause mortality (adjusted for age, sex, comorbidities, and previous cardiovascular events) was significantly increased in patients with possible autonomous cortisol secretion (HR 1·52, 95% CI 1·19–1·94) and autonomous cortisol secretion (1·77, 1·20–2·62) compared with patients with non-functioning adenoma. In women younger than 65 years, autonomous cortisol secretion was associated with higher all-cause mortality than non-functioning adenoma (HR 4·39, 95% CI 1·93–9·96), although this was not observed in men. Cardiometabolic comorbidities were significantly less frequent with non-functioning adenoma than with possible autonomous cortisol secretion and autonomous cortisol secretion (hypertension occurred in 1186 [58·6%] of 2024 patients with non-functioning adenoma, 944 [74·0%] of 1275 with possible autonomous cortisol secretion, and 179 [75·2%] of 238 with autonomous cortisol secretion; dyslipidaemia occurred in 724 [36·2%] of 1999 patients, 547 [43·8%] of 1250, and 123 [51·9%] of 237; and any diabetes occurred in 365 [18·2%] of 2002, 288 [23·0%] of 1250, and 62 [26·7%] of 232; all p values <0·001). Interpretation: Cortisol autonomy is associated with increased all-cause mortality, particularly in women younger than 65 years. However, until results from randomised interventional trials are available, a conservative therapeutic approach seems to be justified in most patients with adrenal incidentaloma. Funding: Deutsche Forschungsgemeinschaft, Associazione Italiana per la Ricerca sul Cancro, Università di Torino

Drug-induced trafficking of p-glycoprotein in human brain capillary endothelial cells as demonstrated by exposure to mitomycin C.

P-glycoprotein (Pgp; ABCB1/MDR1) is a major efflux transporter at the blood-brain barrier (BBB), restricting the penetration of various compounds. In other tissues, trafficking of Pgp from subcellular stores to the cell surface has been demonstrated and may constitute a rapid way of the cell to respond to toxic compounds by functional membrane insertion of the transporter. It is not known whether drug-induced Pgp trafficking also occurs in brain capillary endothelial cells that form the BBB. In this study, trafficking of Pgp was investigated in human brain capillary endothelial cells (hCMEC/D3) that were stably transfected with a doxycycline-inducible MDR1-EGFP fusion plasmid. In the presence of doxycycline, these cells exhibited a 15-fold increase in Pgp-EGFP fusion protein expression, which was associated with an increased efflux of the Pgp substrate rhodamine 123 (Rho123). The chemotherapeutic agent mitomycin C (MMC) was used to study drug-induced trafficking of Pgp. Confocal fluorescence microscopy of single hCMEC/D3-MDR1-EGFP cells revealed that Pgp redistribution from intracellular pools to the cell surface occurred within 2 h of MMC exposure. Pgp-EGFP exhibited a punctuate pattern at the cell surface compatible with concentrated regions of the fusion protein in membrane microdomains, i.e., lipid rafts, which was confirmed by Western blot analysis of biotinylated cell surface proteins in Lubrol-resistant membranes. MMC exposure also increased the functionality of Pgp as assessed in three functional assays with Pgp substrates (Rho123, eFluxx-ID Gold, calcein-AM). However, this increase occurred with some delay after the increased Pgp expression and coincided with the release of Pgp from the Lubrol-resistant membrane complexes. Disrupting rafts by depleting the membrane of cholesterol increased the functionality of Pgp. Our data present the first direct evidence of drug-induced Pgp trafficking at the human BBB and indicate that Pgp has to be released from lipid rafts to gain its full functionality

Identification of molecular pathways affected by pterostilbene, a natural dimethylether analog of resveratrol

<p>Abstract</p> <p>Background</p> <p>Pterostilbene, a naturally occurring phenolic compound produced by agronomically important plant genera such as <it>Vitis </it>and <it>Vacciunium</it>, is a phytoalexin exhibiting potent antifungal activity. Additionally, recent studies have demonstrated several important pharmacological properties associated with pterostilbene. Despite this, a systematic study of the effects of pterostilbene on eukaryotic cells at the molecular level has not been previously reported. Thus, the aim of the present study was to identify the cellular pathways affected by pterostilbene by performing transcript profiling studies, employing the model yeast <it>Saccharomyces cerevisiae</it>.</p> <p>Methods</p> <p><it>S. cerevisiae </it>strain S288C was exposed to pterostilbene at the IC₅₀concentration (70 μM) for one generation (3 h). Transcript profiling experiments were performed on three biological replicate samples using the Affymetrix GeneChip Yeast Genome S98 Array. The data were analyzed using the statistical methods available in the GeneSifter microarray data analysis system. To validate the results, eleven differentially expressed genes were further examined by quantitative real-time RT-PCR, and <it>S. cerevisiae </it>mutant strains with deletions in these genes were analyzed for altered sensitivity to pterostilbene.</p> <p>Results</p> <p>Transcript profiling studies revealed that pterostilbene exposure significantly down-regulated the expression of genes involved in methionine metabolism, while the expression of genes involved in mitochondrial functions, drug detoxification, and transcription factor activity were significantly up-regulated. Additional analyses revealed that a large number of genes involved in lipid metabolism were also affected by pterostilbene treatment.</p> <p>Conclusion</p> <p>Using transcript profiling, we have identified the cellular pathways targeted by pterostilbene, an analog of resveratrol. The observed response in lipid metabolism genes is consistent with its known hypolipidemic properties, and the induction of mitochondrial genes is consistent with its demonstrated role in apoptosis in human cancer cell lines. Furthermore, our data show that pterostilbene has a significant effect on methionine metabolism, a previously unreported effect for this compound.</p