Sensory photoreceptor proteins underpin light-dependent adaptations in nature and enable the optogenetic control of organismal behavior and physiology. We identified the bacterial ...light-oxygen-voltage (LOV) photoreceptor PAL that sequence-specifically binds short RNA stem loops with around 20 nM affinity in blue light and weaker than 1 µM in darkness. A crystal structure rationalizes the unusual receptor architecture of PAL with C-terminal LOV photosensor and N-terminal effector units. The light-activated PAL-RNA interaction can be harnessed to regulate gene expression at the RNA level as a function of light in both bacteria and mammalian cells. The present results elucidate a new signal-transduction paradigm in LOV receptors and conjoin RNA biology with optogenetic regulation, thereby paving the way toward hitherto inaccessible optoribogenetic modalities.
MS-based proteomics was applied to the analysis of the medicinal plant Artemisia annua, exploiting a recently published contig sequence database (Graham et al. (2010) Science 327, 328-331) and other ...genomic and proteomic sequence databases for comparison. A. annua is the predominant natural source of artemisinin, the precursor for artemisinin-based combination therapies (ACTs), which are the WHO-recommended treatment for P. falciparum malaria.
The comparison of various databases containing A. annua sequences (NCBInr/viridiplantae, UniProt/viridiplantae, UniProt/A. annua, an A. annua trichome Trinity contig database, the above contig database and another A. annua EST database) revealed significant differences in respect of their suitability for proteomic analysis, showing that an organism-specific database that has undergone extensive curation, leading to longer contig sequences, can greatly increase the number of true positive protein identifications, while reducing the number of false positives. Compared to previously published data an order-of-magnitude more proteins have been identified from trichome-enriched A. annua samples, including proteins which are known to be involved in the biosynthesis of artemisinin, as well as other highly abundant proteins, which suggest additional enzymatic processes occurring within the trichomes that are important for the biosynthesis of artemisinin.
The newly gained information allows for the possibility of an enzymatic pathway, utilizing peroxidases, for the less well understood final stages of artemisinin's biosynthesis, as an alternative to the known non-enzymatic in vitro conversion of dihydroartemisinic acid to artemisinin. Data are available via ProteomeXchange with identifier PXD000703.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The organocatalytic transformation of resorcinols is extremely rare. In this article, we report a highly enantioselective, organocatalytic intramolecular cyclization of these systems by a ...Friedel–Crafts‐type 1,4‐addition using a Jørgensen‐Hayashi‐like organocatalyst with a large silyl protecting group, and show that heat improves reaction yield with virtually no detriment to enantioselectivity. A variety of bicyclic resorcinols were obtained with excellent enantioselectivities (up to 94%). To show the utility of these constructs, and as part of a wider project involving the synthesis of cannabinoid‐like compounds, the resorcinol formed was used to generate both ‘normal’ and ‘abnormal’ cannabidiol (CBD) derivatives which were shown to have anticonvulsant activity.
Cinchona alkaloids with a free 6'-OH functionality are being increasingly used within asymmetric organocatalysis. This fascinating class of bifunctional catalyst offers a genuine alternative to the ...more commonly used thiourea systems and because of the different spacing between the functional groups, can control enantioselectivity where other organocatalysts have failed. In the main, this review covers the highlights from the last five years and attempts to show the diversity of reactions that these systems can control. It is hoped that chemists developing asymmetric methodologies will see the value in adding these easily accessible, but underused organocatalysts to their screens.
Clickmers are chemically modified aptamers representing an innovative reagent class for developing binders for biomolecules with great impact on therapeutic and diagnostic applications. To establish ...a novel layer for screening various chemical entities, we developed a split-combine selection strategy simultaneously enriching for clickmers having different modifications. Due to the inherent design of this strategy, dynamic changes of DNA populations are traceable at an individual sequence level. Besides off-rate guided enrichment, the process makes the survival of the sequences most adapted to the applied selection condition observable. The underlying strategy provides unprecedented molecular insight into the selection process, based on which more sophisticated procedures will become pliable in the future.
A split-combine selection approach reveals dynamic population changes in DNA libraries during
in vitro
selection procedures.
OBJECTIVE:This study aimed to a) compare the efficacy of metoclopramide and erythromycin in the treatment of feed intolerance in critical illness; and b) determine the effectiveness of “rescue” ...combination therapy in patients who fail monotherapy.
DESIGN:Randomized controlled trial.
SETTING:Level III mixed medical and surgical intensive care unit.
PATIENTS:Ninety mechanically ventilated, medical patients with feed-intolerance (gastric residual volume ≥250 mL).
INTERVENTIONS:Patients received either metoclopramide 10 mg intravenously four times daily (n = 45) or erythromycin 200 mg intravenously twice a day (n = 45) in a double-blind, randomized fashion. After the first dose, nasogastric feeding was commenced and 6-hourly nasogastric aspirates were performed. If a gastric residual volume ≥250 mL recurred on treatment, open-label, combination therapy was given. Patients were studied for 7 days. Successful feeding was defined as 6-hourly gastric residual volume <250 mL with a feeding rate ≥40 mL/hr.
MEASUREMENTS AND MAIN RESULTS:Demographic data, blood glucose levels, and use of inotropes, opioids, and benzodiazepines were similar between the two groups. After 24 hrs of treatment, both monotherapies reduced the mean gastric residual volume (metoclopramide, 830 ± 32 mL to 435 ± 30 mL, p < .0001; erythromycin, 798 ± 33 mL to 201 ± 19 mL, p < .0001) and improved the proportion of patients with successful feeding (metoclopramide = 62% and erythromycin = 87%). Treatment with erythromycin was more effective than metoclopramide, but the effectiveness of both treatments declined rapidly over time. In patients who failed monotherapy, rescue combination therapy was highly effective (day 1 = 92%) and maintained its effectiveness for the study duration (day 6 = 67%). High pretreatment gastric residual volume was associated with poor response to prokinetic therapy.
CONCLUSIONS:In critical illness, erythromycin is more effective than metoclopramide in treating feed intolerance, but the rapid decline in effectiveness renders both treatments suboptimal. Rescue combination therapy is highly effective, and further study is required to examine its role as the first-line therapy.
OBJECTIVE:To compare the efficacy of combination therapy, with erythromycin and metoclopramide, to erythromycin alone in the treatment of feed intolerance in critically ill patients.
...DESIGN:Randomized, controlled, double-blind trial.
SETTING:Mixed medical and surgical intensive care unit.
PATIENTS:Seventy-five mechanically ventilated, medical patients with feed intolerance (gastric residual volume ≥250 mL).
INTERVENTIONS:Patients received either combination therapy (n = 37; 200 mg of intravenous erythromycin twice daily + 10 mg of intravenous metoclopramide four times daily) or erythromycin alone (n = 38; 200 mg of intravenous erythromycin twice daily) in a prospective, randomized fashion. Gastric feeding was re-commenced and 6-hourly gastric aspirates performed. Patients were studied for 7 days. Successful feeding was defined as a gastric residual volume <250 mL with the feeding rate ≥40 mL/hr, over 7 days. Secondary outcomes included daily caloric intake, vomiting, postpyloric feeding, length of stay, and mortality.
MEASUREMENTS AND MAIN RESULTS:Demographic data; use of inotropes, opioids, or benzodiazepines; and pretreatment gastric residual volume were similar between the two groups. The gastric residual volume was significantly lower after 24 hrs of treatment with combination therapy, compared with erythromycin alone (136 ± 23 mL vs. 293 ± 45 mL, p = .04). Over the 7 days, patients treated with combination therapy had greater feeding success, received more daily calories, and had a lower requirement for postpyloric feeding, compared with erythromycin alone. Tachyphylaxis occurred in both groups but was less with combination therapy. Sedation, higher pretreatment gastric residual volume, and hypoalbuminemia were significantly associated with a poor response. There was no difference in the length of hospital stay or mortality rate between the groups. Watery diarrhea was more common with combination therapy (20 of 37 vs. 10 of 38, p = .01) but was not associated with enteric infections, including Clostridium difficile.
CONCLUSIONS:In critically ill patients with feed intolerance, combination therapy with erythromycin and metoclopramide is more effective than erythromycin alone in improving the delivery of nasogastric nutrition and should be considered as the first-line treatment.
Abstract
Background
Defining epithelial cell contributions to inflammatory bowel disease (IBD) is essential for the development of much needed therapies for barrier repair. Children with very early ...onset (VEO)-IBD have more extensive, severe, and refractory disease than older children and adults with IBD and, in some cases, have defective barrier function. We therefore evaluated functional and transcriptomic differences between pediatric IBD (VEO and older onset) and non-IBD epithelium using 3-dimensional, biopsy-derived organoids.
Methods
We measured growth efficiency relative to histopathological and clinical parameters in patient enteroid (ileum) and colonoid (colon) lines. We performed RNA-sequencing on patient colonoids and subsequent flow cytometry after multiple passages to evaluate changes that persisted in culture.
Results
Enteroids and colonoids from pediatric patients with IBD exhibited decreased growth associated with histological inflammation compared with non-IBD controls. We observed increased LYZ expression in colonoids from pediatric IBD patients, which has been reported previously in adult patients with IBD. We also observed upregulation of antigen presentation genes HLA-DRB1 and HLA-DRA, which persisted after prolonged passaging in patients with pediatric IBD.
Conclusions
We present the first functional evaluation of enteroids and colonoids from patients with VEO-IBD and older onset pediatric IBD, a subset of which exhibits poor growth. Enhanced, persistent epithelial antigen presentation gene expression in patient colonoids supports the notion that epithelial cell-intrinsic differences may contribute to IBD pathogenesis.
Graphical Abstract
Graphical Abstract
Malnutrition is associated with poor outcomes in critically ill patients. Although nutritional support is yet to be proven to improve mortality in non-malnourished critically ill patients, early ...enteral feeding is considered best practice. However, enteral feeding is often limited by delayed gastric emptying. The best method to clinically identify delayed gastric emptying and feed intolerance is unclear. Gastric residual volume (GRV) measured at the bedside is widely used as a surrogate marker for gastric emptying, but the value of GRV measurement has recently been disputed. While the mechanisms underlying delayed gastric emptying require further investigation, recent research has given a better appreciation of the pathophysiology. A number of pharmacological strategies are available to improve the success of feeding. Recent data suggest a combination of intravenous metoclopramide and erythromycin to be the most successful treatment, but novel drug therapies should be explored. Simpler methods to access the duodenum and more distal small bowel for feed delivery are also under investigation. This review summarises current understanding of the factors responsible for, and mechanisms underlying feed intolerance in critical illness, together with the evidence for current practices. Areas requiring further research are also highlighted.
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