-
Microbial Cell Factories Aug 2021Melatonin has attracted substantial attention because of its excellent prospects for both medical applications and crop improvement. The microbial production of...
BACKGROUND
Melatonin has attracted substantial attention because of its excellent prospects for both medical applications and crop improvement. The microbial production of melatonin is a safer and more promising alternative to chemical synthesis approaches. Researchers have failed to produce high yields of melatonin in common heterologous hosts due to either the insolubility or low enzyme activity of proteins encoded by gene clusters related to melatonin biosynthesis.
RESULTS
Here, a combinatorial gene pathway for melatonin production was successfully established in Escherichia coli by combining the physostigmine biosynthetic genes from Streptomyces albulus and gene encoding phenylalanine 4-hydroxylase (P4H) from Xanthomonas campestris and caffeic acid 3-O-methyltransferase (COMT) from Oryza sativa. A threefold improvement of melatonin production was achieved by balancing the expression of heterologous proteins and adding 3% glycerol. Further protein engineering and metabolic engineering were conducted to improve the conversion of N-acetylserotonin (NAS) to melatonin. Construction of COMT variant containing C303F and V321T mutations increased the production of melatonin by fivefold. Moreover, the deletion of speD gene increased the supply of S-adenosylmethionine (SAM), an indispensable cofactor of COMT, which doubled the yield of melatonin. In the final engineered strain EcMEL8, the production of NAS and melatonin reached 879.38 ± 71.42 mg/L and 136.17 ± 1.33 mg/L in a shake flask. Finally, in a 2-L bioreactor, EcMEL8 produced 1.06 ± 0.07 g/L NAS and 0.65 ± 0.11 g/L melatonin with tryptophan supplementation.
CONCLUSIONS
This study established a novel combinatorial pathway for melatonin biosynthesis in E. coli and provided alternative strategies for improvement of melatonin production.
Topics: Escherichia coli; Melatonin; Metabolic Engineering; Protein Engineering
PubMed: 34454478
DOI: 10.1186/s12934-021-01662-8 -
Angewandte Chemie (International Ed. in... Oct 2021Enantioselective synthesis of bioactive compounds bearing a pyrroloindole framework is often laborious. In contrast, there are several S-adenosyl methionine...
Enantioselective synthesis of bioactive compounds bearing a pyrroloindole framework is often laborious. In contrast, there are several S-adenosyl methionine (SAM)-dependent methyl transferases known for stereo- and regioselective methylation at the C3 position of various indoles, directly leading to the formation of the desired pyrroloindole moiety. Herein, the SAM-dependent methyl transferase PsmD from Streptomyces griseofuscus, a key enzyme in the biosynthesis of physostigmine, is characterized in detail. The biochemical properties of PsmD and its substrate scope were demonstrated. Preparative scale enzymatic methylation including SAM regeneration was achieved for three selected substrates after a design-of-experiment optimization.
Topics: Biocatalysis; Indoles; Kinetics; Methylation; Methyltransferases; Pyrroles; S-Adenosylmethionine; Stereoisomerism; Streptomyces
PubMed: 34399441
DOI: 10.1002/anie.202107619 -
Neurogastroenterology and Motility Mar 2022This systematic review and meta-analysis aimed to evaluate the effects of pharmacological agents for neurogenic oropharyngeal dysphagia based on evidence from randomized... (Meta-Analysis)
Meta-Analysis
BACKGROUND
This systematic review and meta-analysis aimed to evaluate the effects of pharmacological agents for neurogenic oropharyngeal dysphagia based on evidence from randomized controlled trials (RCTs).
METHODS
Electronic databases were systematically searched between January 1970 and March 2021. Two reviewers independently extracted and synthesized the data. The outcome measure was changed in (any) relevant clinical swallowing-related characteristics.
KEY RESULTS
Data from 2186 dysphagic patients were collected from 14 RCT studies across a range of pharmacotherapies. The pooled effect size of transient receptor potential (TRP) channel agonists was large compared to placebo interventions (SMD[95%CI] =1.27[0.74,1.80], p < 0.001; I = 79%). Data were limited for other pharmacological agents and the overall pooled effect size of these agents was non-significant (SMD [95% CI] =0.25 [-0.24, 0.73]; p = 0.31; I = 85%). When analyzed separately, large effect sizes were observed with Nifedipine (SMD[95%CI] =1.13[0.09,2.18]; p = 0.03) and Metoclopramide (SMD[95%CI] =1.68[1.08,2.27]; p < 0.001). By contrast, the effects of angiotensin-converting enzyme (ACE) inhibitors (SMD[95%CI] = -0.67[-2.32,0.99]; p = 0.43; I = 61%), Physostigmine (SMD[95%CI] = -0.05[-1.03,0.93]; p = 0.92) and Glyceryl Trinitrate (GTN) (SMD [95% CI] = -0.01 [-0.11, 0.08]; p = 0.78) were non-significant. Within stroke patients, subgroup analysis showed that TRP channel agonists had a moderate pooled effect size (SMD[95%CI] =0.74[0.10,1.39]; p = 0.02; I = 82%) whereas the effects of other agents were non-significant (SMD[95%CI] =0.40[-0.04,0.84]; p = 0.07; I = 87%).
CONCLUSIONS & INFERENCES
Our results showed that TRP channel agonists, Nifedipine and Metoclopromide may be beneficial for neurogenic dysphagic patients. Large scale, multicenter clinical trials are warranted to fully explore their therapeutic effects on swallowing.
Topics: Deglutition; Deglutition Disorders; Humans; Multicenter Studies as Topic; Nifedipine; Stroke
PubMed: 34337829
DOI: 10.1111/nmo.14220 -
Neuroscience Insights 2021Among the various chemicals that are commonly used as pesticides, organophosphates (OPs), and to a lesser extent, carbamates, are most frequently associated with adverse...
Among the various chemicals that are commonly used as pesticides, organophosphates (OPs), and to a lesser extent, carbamates, are most frequently associated with adverse long-term neurological consequences. OPs and the carbamate, pyridostigmine, used as a prophylactic drug against potential nerve agent attacks, have also been implicated in Gulf War Illness (GWI), which is often characterized by chronic neurological symptoms. While most OP- and carbamate-based pesticides, and pyridostigmine are relatively potent acetylcholinesterase inhibitors (AChEIs), this toxicological mechanism is inadequate to explain their long-term health effects, especially when no signs of acute cholinergic toxicity are exhibited. Our previous work suggests that a potential mechanism of the long-term neurological deficits associated with OPs is impairment of axonal transport (AXT); however, we had not previously evaluated carbamates for this effect. Here we thus evaluated the carbamate, physostigmine (PHY), a highly potent AChEI, on AXT using an neuronal live imaging assay that we have previously found to be very sensitive to OP-related deficits in AXT. We first evaluated the OP, diisopropylfluorophosphate (DFP) (concentration range 0.001-10.0 µM) as a reference compound that we found previously to impair AXT and subsequently evaluated PHY (concentration range 0.01-100 nM). As expected, DFP impaired AXT in a concentration-dependent manner, replicating our previously published results. In contrast, none of the concentrations of PHY (including concentrations well above the threshold for impairing AChE) impaired AXT. These data suggest that the long-term neurological deficits associated with some carbamates are not likely due to acute impairments of AXT.
PubMed: 34104889
DOI: 10.1177/26331055211020289 -
Behavioural Brain Research Jul 2021There is high clinical interest in improving the pharmacological treatment of individuals with Major Depressive Disorder (MDD). This neuropsychiatric disorder continues...
There is high clinical interest in improving the pharmacological treatment of individuals with Major Depressive Disorder (MDD). This neuropsychiatric disorder continues to cause significant morbidity and mortality worldwide, where existing pharmaceutical treatments such as selective serotonin reuptake inhibitors often have limited efficacy. In a recent publication, we demonstrated an antidepressant-like role for the acetylcholinesterase inhibitor (AChEI) donepezil in the C57BL/6J mouse forced swim test (FST). Those data added to a limited literature in rodents and human subjects which suggests AChEIs have antidepressant properties, but added the novel finding that donepezil only showed antidepressant-like properties at lower doses (0.02, 0.2 mg/kg). At a high dose (2.0 mg/kg), donepezil tended to promote depression-like behavior, suggesting a u-shaped dose-response curve for FST immobility. Here we investigate the effects of three other AChEIs with varying molecular structures: galantamine, physostigmine, and rivastigmine, to test whether they also exhibit antidepressant-like effects in the FST. We find that these drugs do exhibit therapeutic-like effects at low but not high doses, albeit at lower doses for physostigmine. Further, we find that their antidepressant-like effects are not mediated by generalized hyperactivity in the novel open field test, and are also not accompanied by anxiolytic-like properties. These data further support the hypothesis that acetylcholine has a u-shaped dose-response relationship with immobility in the C57BL/6J mouse FST, and provide a rationale for more thoroughly investigating whether reversible AChEIs as a class can be repurposed for the treatment of MDD in human subjects.
Topics: Animals; Antidepressive Agents; Association Learning; Behavior, Animal; Cholinesterase Inhibitors; Donepezil; Dose-Response Relationship, Drug; Drug Repositioning; Galantamine; Male; Mice; Mice, Inbred C57BL; Motor Activity; Physostigmine; Rivastigmine; Swimming
PubMed: 33910028
DOI: 10.1016/j.bbr.2021.113323 -
British Journal of Clinical Pharmacology Nov 2021HTL0009936 is a selective M muscarinic receptor agonist in development for cognitive dysfunction in Alzheimer's disease. Safety, tolerability and pharmacokinetics and... (Randomized Controlled Trial)
Randomized Controlled Trial
AIMS
HTL0009936 is a selective M muscarinic receptor agonist in development for cognitive dysfunction in Alzheimer's disease. Safety, tolerability and pharmacokinetics and exploratory pharmacodynamic effects of HTL0009936 administered by continuous IV infusion at steady state were investigated in elderly subjects with below average cognitive functioning (BACF).
METHODS
Part A was a four-treatment open label sequential study in healthy elderly investigating 10-83 mg HTL0009936 (IV) and a 24 mg HTL0009936 single oral dose. Part B was a five-treatment randomized, double-blind, placebo and physostigmine controlled cross-over study with IV HTL0009936 in elderly subjects with BACF. Pharmacodynamic assessments were performed using neurocognitive and electrophysiological tests.
RESULTS
Pharmacokinetics of HTL0009936 showed dose-proportional increases in exposure with a mean half-life of 2.4 hours. HTL0009936 was well-tolerated with transient dose-related adverse events (AEs). Small increases in mean systolic blood pressure of 7.12 mmHg (95% CI [3.99-10.24]) and in diastolic of 5.32 mmHg (95% CI [3.18-7.47]) were noted at the highest dose in part B. Overall, there was suggestive, but no definitive, positive or negative pharmacodynamic effects. Statistically significant effects were observed on P300 with HTL0009936 and adaptive tracking with physostigmine.
CONCLUSIONS
HTL0009936 showed well-characterized pharmacokinetics and single doses were safe and generally well-tolerated in healthy elderly subjects. Due to physostigmine tolerability issues and subject burden, the study design was changed and some pharmacodynamic assessments (neurocognitive) were performed at suboptimal drug exposures. Therefore no clear conclusions can be made on pharmacodynamic effects of HTL0009936, although an effect on P300 is suggestive of central target engagement.
Topics: Aged; Area Under Curve; Cholinergic Agents; Cross-Over Studies; Dose-Response Relationship, Drug; Double-Blind Method; Humans; Receptors, Cholinergic
PubMed: 33891333
DOI: 10.1111/bcp.14872 -
Journal of Experimental Botany Jun 2021Plant association with arbuscular mycorrhizal fungi (AMF) can increase their ability to overcome multiple stresses, but their impact on plant interactions with...
Plant association with arbuscular mycorrhizal fungi (AMF) can increase their ability to overcome multiple stresses, but their impact on plant interactions with herbivorous insects is controversial. Here we show higher mortality of the leaf-chewer Spodoptera exigua when fed on tomato plants colonized by the AMF Funneliformis mosseae, evidencing mycorrhiza-induced resistance. In search of the underlying mechanisms, an untargeted metabolomic analysis through ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS) was performed. The results showed that mycorrhizal symbiosis had a very limited impact on the leaf metabolome in the absence of stress, but significantly modulated the response to herbivory in the damaged area. A cluster of over accumulated metabolites was identified in those leaflets damaged by S. exigua feeding in mycorrhizal plants, while unwounded distal leaflets responded similar to those from non-mycorrhizal plants. These primed-compounds were mostly related to alkaloids, fatty acid derivatives and phenylpropanoid-polyamine conjugates. The deleterious effect on larval survival of some of these compounds, including the alkaloid physostigmine, the fatty acid derivatives 4-oxododecanedioic acid and azelaic acid, was confirmed. Thus, our results evidence the impact of AMF on metabolic reprograming upon herbivory that leads to a primed accumulation of defensive compounds.
Topics: Animals; Chromatography, Liquid; Fungi; Herbivory; Solanum lycopersicum; Mycorrhizae; Symbiosis; Tandem Mass Spectrometry
PubMed: 33884424
DOI: 10.1093/jxb/erab171 -
Cureus Mar 2021Anticholinergic toxicity is a relatively uncommon occurrence. When it does occur, it is usually attributed to an overdose of anticholinergic agents, especially in the...
Anticholinergic toxicity is a relatively uncommon occurrence. When it does occur, it is usually attributed to an overdose of anticholinergic agents, especially in the elderly population. In younger patients, anticholingeric toxicity is usually due to an intentional overdose in a suicide attempt, accidental exposure to jimson weed or deadly nightshade plant, or the combination of anticholinergic drugs with recreational drugs (psilocybin mushroom). Over-the-counter cold medicines are well known to contain a variety of anticholinergic substances, the most well-known being diphenhydramine. Uncommonly, these readily available substances can produce anticholinergic toxicity in elderly patients, even when appropriate dosages are consumed. Younger patients are less likely to develop this clinical picture, due to higher renal clearance and lower drug volume of distribution. Nonetheless, clinical suspicion should remain high in the younger population. Patients can present with fever, tachycardia, diplopia, urinary retention, dry mucous membranes, and altered mental status. This case provides awareness to the unlikely complication of over-the-counter cold medicines in a young 19-year-old patient, while highlighting the need for diligent history taking and deliberate use of physostigmine in patients with altered mental status.
PubMed: 33880270
DOI: 10.7759/cureus.13919 -
The American Journal of Case Reports Feb 2021BACKGROUND Clozapine is a well-proven atypical antipsychotic drug used for therapy of treatment-resistant schizophrenia. Over the last decades only a few cases of...
BACKGROUND Clozapine is a well-proven atypical antipsychotic drug used for therapy of treatment-resistant schizophrenia. Over the last decades only a few cases of clozapine poisoning have been reported. Hence, guidelines for in-hospital management are currently not available. Most of the reported cases underwent detoxication measures as charcoal therapy and/or gastric lavage. However, there is no evidence for primary detoxication to improve clinical outcome. In contrast, use of therapy with intravenous physostigmine in the case of anticholinergic syndrome is restricted due to concerns about safety and dosing. We present a case of acute high-dose clozapine poisoning without detoxication and complete recovery supported by physostigmine. CASE REPORT We report the case of a 28-year-old man with prior diagnosed schizophrenia who presumably ingested 8 g (regular maximum daily dose 900 mg/d) of clozapine with uncertain intent. Initial computed tomography (CT) showed pulmonary infiltrates and widespread pneumomediastinum and soft-tissue emphysema of unknown genesis. The patient developed a progressive impairment of vigilance and respiratory insufficiency requiring invasive artificial ventilation for 31 h. Afterwards, an anticholinergic syndrome led again to impaired vigilance, tachycardia, and hyperventilation. To avoid risks associated with artificial ventilation, we applied physostigmine. Subsequently, the anticholinergic syndrome and the pneumomediastinum completely regressed and no further artificial ventilation was needed. CONCLUSIONS Based on the presumably ingested dosage, we present the likely highest reported nonfatal overdose of clozapine without detoxication. Additionally, we observed widespread pneumomediastinum as an uncommon complication. Our approach was to refrain from detoxication to minimize complications and to treat early with physostigmine because of anticholinergic syndrome to minimize its impact and to avoid artificial ventilation due do vigilance impairment.
Topics: Adult; Antipsychotic Agents; Clozapine; Gastric Lavage; Humans; Male; Physostigmine; Schizophrenia
PubMed: 33591960
DOI: 10.12659/AJCR.929147 -
Frontiers in Behavioral Neuroscience 2020When stress becomes chronic it can trigger lasting brain and behavioral changes including Major Depressive Disorder (MDD). There is conflicting evidence regarding... (Review)
Review
When stress becomes chronic it can trigger lasting brain and behavioral changes including Major Depressive Disorder (MDD). There is conflicting evidence regarding whether acetylcholinesterase inhibitors (AChEIs) may have antidepressant properties. In a recent publication, we demonstrated a strong dose-dependency of the effect of AChEIs on antidepressant-related behavior in the mouse forced swim test: whereas the AChEI donepezil indeed promotes depression-like behavior at a high dose, it has antidepressant-like properties at lower doses in the same experiment. Our data therefore suggest a Janus-faced dose-response curve for donepezil in depression-related behavior. In this review, we investigate the mood-related properties of AChEIs in greater detail, focusing on both human and rodent studies. In fact, while there have been many studies showing pro-depressant activity by AChEIs and this is a major concept in the field, a variety of other studies in both humans and rodents show antidepressant effects. Our study was one of the first to systematically vary dose to include very low concentrations while measuring behavioral effects, potentially explaining the apparent disparate findings in the field. The possibility of antidepressant roles for AChEIs in rodents may provide hope for new depression treatments. Importantly, MDD is a psychosocial stress-linked disorder, and in rodents, stress is a major experimental manipulation for studying depression mechanisms, so an important future direction will be to determine the extent to which these depression-related effects are stress-sensitive. In sum, gaining a greater understanding of the potentially therapeutic mood-related effects of low dose AChEIs, both in rodent models and in human subjects, should be a prioritized topic in ongoing translational research.
PubMed: 33519395
DOI: 10.3389/fnbeh.2020.620119