Your browser doesn't support javascript.
Show: 20 | 50 | 100
Results 1 - 20 de 7.135
Filter
1.
Pediatr Int ; 64(1): e15153, 2022 Jan.
Article in English | MEDLINE | ID: mdl-35522644

ABSTRACT

BACKGROUND: We investigated the efficacy of sivelestat sodium hydrate (SSH) as a treatment for Kawasaki disease, and its pharmacological action sites, in mice with Candida albicans water-soluble fraction-induced vasculitis. METHODS: Sivelestat sodium hydrate was administered intraperitoneally to Candida albicans water-soluble fraction-induced vasculitis model mice to assess its efficacy in preventing the development of coronary artery lesions based on the degree of inflammatory cell infiltration in the aortic root and coronary arteries (vasculitis score). The pharmacological sites of action were investigated based on changes in neutrophil elastase (NE) and intercellular adhesion molecule 1 (ICAM-1) positive areas, ICAM-1 and tumor necrosis factor-α mRNA expression levels in the upper heart, and the proportion of monocytes in the peripheral blood. RESULTS: The vasculitis score decreased below the lower limit of the 95% confidence interval of untreated mice in 69% of the SSH-treated mice. The NE- and ICAM-1-positive regions, and the mRNA expression of ICAM-1 and tumor necrosis factor-α were lower in the SSH-treated mice than in the untreated mice. The proportion of monocytes in the peripheral blood was higher in the SSH-treated mice than in the untreated mice, whereas monocyte migration to inflammation areas was suppressed in the SSH-treated mice. CONCLUSIONS: Our results showed that SSH might prevent the development of coronary artery lesions and ameliorate disease activity. In addition to its NE-inhibitory effect, SSH sites of action may also include monocytes.


Subject(s)
Glycine , Mucocutaneous Lymph Node Syndrome , Sulfonamides , Vasculitis , Animals , Candida albicans , Glycine/analogs & derivatives , Glycine/pharmacology , Humans , Intercellular Adhesion Molecule-1/genetics , Mice , Mucocutaneous Lymph Node Syndrome/drug therapy , RNA, Messenger , Sulfonamides/pharmacology , Tumor Necrosis Factor-alpha , Vasculitis/chemically induced , Vasculitis/drug therapy
2.
Cells ; 11(8)2022 Apr 15.
Article in English | MEDLINE | ID: mdl-35456035

ABSTRACT

Mechanical allodynia (pain to normally innocuous tactile stimuli) is a widespread symptom of inflammatory and neuropathic pain. Spinal or medullary dorsal horn (SDH or MDH) circuits mediating tactile sensation and pain need to interact in order to evoke mechanical allodynia. PKCγ-expressing (PKCγ+) interneurons and inhibitory controls within SDH/MDH inner lamina II (IIi) are pivotal in connecting touch and pain circuits. However, the relative contribution of GABA and glycine to PKCγ+ interneuron inhibition remains unknown. We characterized inhibitory inputs onto PKCγ+ interneurons by combining electrophysiology to record spontaneous and miniature IPSCs (sIPSCs, mIPSCs) and immunohistochemical detection of GABAARα2 and GlyRα1 subunits in adult rat MDH. While GlyR-only- and GABAAR-only-mediated mIPSCs/sIPSCs are predominantly recorded from PKCγ+ interneurons, immunohistochemistry reveals that ~80% of their inhibitory synapses possess both GABAARα2 and GlyRα1. Moreover, nearly all inhibitory boutons at gephyrin-expressing synapses on these cells contain glutamate decarboxylase and are therefore GABAergic, with around half possessing the neuronal glycine transporter (GlyT2) and therefore being glycinergic. Thus, while GABA and glycine are presumably co-released and GABAARs and GlyRs are present at most inhibitory synapses on PKCγ+ interneurons, these interneurons exhibit almost exclusively GABAAR-only and GlyR-only quantal postsynaptic inhibitory currents, suggesting a pharmacological specialization of their inhibitory synapses.


Subject(s)
Hyperalgesia , Receptors, Glycine , Animals , Glycine/pharmacology , Interneurons/metabolism , Pain , Rats , Receptors, Glycine/metabolism , Substantia Gelatinosa/metabolism , Synaptic Transmission/physiology , gamma-Aminobutyric Acid
3.
Redox Biol ; 52: 102313, 2022 06.
Article in English | MEDLINE | ID: mdl-35447412

ABSTRACT

Lower circulating levels of glycine are consistently reported in association with cardiovascular disease (CVD), but the causative role and therapeutic potential of glycine in atherosclerosis, the underlying cause of most CVDs, remain to be established. Here, following the identification of reduced circulating glycine in patients with significant coronary artery disease (sCAD), we investigated a causative role of glycine in atherosclerosis by modulating glycine availability in atheroprone mice. We further evaluated the atheroprotective potential of DT-109, a recently identified glycine-based compound with dual lipid/glucose-lowering properties. Glycine deficiency enhanced, while glycine supplementation attenuated, atherosclerosis development in apolipoprotein E-deficient (Apoe-/-) mice. DT-109 treatment showed the most significant atheroprotective effects and lowered atherosclerosis in the whole aortic tree and aortic sinus concomitant with reduced superoxide. In Apoe-/- mice with established atherosclerosis, DT-109 treatment significantly reduced atherosclerosis and aortic superoxide independent of lipid-lowering effects. Targeted metabolomics and kinetics studies revealed that DT-109 induces glutathione formation in mononuclear cells. In bone marrow-derived macrophages (BMDMs), glycine and DT-109 attenuated superoxide formation induced by glycine deficiency. This was abolished in BMDMs from glutamate-cysteine ligase modifier subunit-deficient (Gclm-/-) mice in which glutathione biosynthesis is impaired. Metabolic flux and carbon tracing experiments revealed that glycine deficiency inhibits glutathione formation in BMDMs while glycine-based treatment induces de novo glutathione biosynthesis. Through a combination of studies in patients with CAD, in vivo studies using atherosclerotic mice and in vitro studies using macrophages, we demonstrated a causative role of glycine in atherosclerosis and identified glycine-based treatment as an approach to mitigate atherosclerosis through antioxidant effects mediated by induction of glutathione biosynthesis.


Subject(s)
Atherosclerosis , Plaque, Atherosclerotic , Animals , Apolipoproteins E/genetics , Atherosclerosis/drug therapy , Atherosclerosis/genetics , Atherosclerosis/metabolism , Disease Models, Animal , Glutamate-Cysteine Ligase , Glutathione/metabolism , Glycine/pharmacology , Glycine/therapeutic use , Humans , Mice , Mice, Inbred C57BL , Mice, Knockout , Plaque, Atherosclerotic/metabolism , Superoxides
4.
Plant Sci ; 318: 111212, 2022 May.
Article in English | MEDLINE | ID: mdl-35351301

ABSTRACT

The objective of this work was to characterize the resistance mechanisms and the primary metabolism of a multiple resistant (MR) population of Amaranthus palmeri to glyphosate and to the acetolactate synthase (ALS) inhibitor pyrithiobac. All MR plants analysed were glyphosate-resistant due to 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene amplification. Resistance to pyrithiobac was more variable among individuals and was related to point mutations at five positions in the ALS gene sequence: A122, A205, W574, S653 and G654. All MR plants were heterozygous for W574, the most abundant mutation. In nontreated plants, the presence of mutations did not affect ALS functionality, and plants with the W574L mutation showed the highest ALS resistance level to pyrithiobac. The accumulation of the transcripts corresponding to several genes of the aromatic amino acid (AAA) and branched-chain amino acid (BCAA) pathways detected in nontreated MR plants indicated additional effects of EPSPS gene amplification and ALS mutations. The physiological performance of the MR population after treatment with glyphosate and/or pyrithiobac was compared with that of a sensitive (S) population. The increase induced in total soluble sugars, AAA or BCAA content by both herbicides was higher in the S population than in the MR population. Physiological effects were not exacerbated after the mixture of both herbicides in S or in MR populations. This study provides new insights into the physiology of a multiple resistant A. palmeri, which could be very useful for achieving effective management of this weed.


Subject(s)
Amaranthus , Herbicides , Amaranthus/genetics , Glycine/analogs & derivatives , Glycine/pharmacology , Herbicide Resistance/genetics , Herbicides/pharmacology
5.
J Neural Transm (Vienna) ; 129(4): 395-407, 2022 Apr.
Article in English | MEDLINE | ID: mdl-35322277

ABSTRACT

Interventions that elevate glycine levels and target the glycine receptor (GlyR) in the nucleus Accumbens (nAc) reduce ethanol intake in rats, supposedly by acting on the brain reward system via increased basal and attenuated ethanol-induced nAc dopamine release. Glycine transport across the blood brain barrier (BBB) appears inefficient, but glycine-containing dipeptides elevate whole brain tissue dopamine levels in mice. This study explores whether treatment with the glycine-containing dipeptides leucine-glycine (Leu-Gly) and glycine-leucine (Gly-Leu) by means of a hypothesized, facilitated BBB passage, alter nAc glycine and dopamine levels and locomotor activity in two rodent models. The acute effects of Leu-Gly and Gly-Leu (1-1000 mg/kg, i.p.) alone or Leu-Gly in combination with ethanol on locomotion in male NMRI mice were examined in locomotor activity boxes. Striatal and brainstem slices were obtained for ex vivo HPLC analyses of tissue levels of glycine and dopamine. Furthermore, the effects of Leu-Gly i.p. (1-1000 mg/kg) on glycine and dopamine output in the nAc were examined using in vivo microdialysis coupled to HPLC in freely moving male Wistar rats. Leu-Gly and Gly-Leu did not significantly alter locomotion, ethanol-induced hyperlocomotor activity or tissue levels of glycine or dopamine, apart from Gly-Leu 10 mg/kg that slightly raised nAc dopamine. Microdialysis revealed no significant alterations in nAc glycine or dopamine levels when regarding all rats as a homogenous group. In a subgroup of rats defined as dopamine responders, a significant elevation of nAc dopamine (20%) was seen following Leu-Gly 10-1000 mg/kg i.p, and this group of animals presented lower baseline dopamine levels compared to dopamine non-responders. To conclude, peripheral injection of glycine-containing dipeptides appears inefficient in elevating central glycine levels but raises accumbal dopamine levels in a subgroup of rats with a lower endogenous dopamine tone. The tentative relationship between dopamine baseline and ensuing response to glycinergic treatment and presumptive direct interactions between glycine-containing dipeptides and the GlyR bear insights for refinement of the glycinergic treatment concept for alcohol use disorder (AUD).


Subject(s)
Dopamine , Glycine , Animals , Dipeptides , Ethanol , Glycine/pharmacology , Leucine , Male , Mice , Microdialysis , Rats , Rats, Wistar , Receptors, Glycine
6.
J Lipid Res ; 63(4): 100192, 2022 Apr.
Article in English | MEDLINE | ID: mdl-35278409

ABSTRACT

Oral and gut Bacteroidetes produce unique classes of serine-glycine lipodipeptides and glycine aminolipids that signal through host Toll-like receptor 2. These glycine lipids have also been detected in human arteries, but their effects on atherosclerosis are unknown. Here, we sought to investigate the bioactivity of bacterial glycine lipids in mouse models of atherosclerosis. Lipid 654 (L654), a serine-glycine lipodipeptide species, was first tested in a high-fat diet (HFD)-fed Ldlr-/- model of atherosclerosis. Intraperitoneal administration of L654 over 7 weeks to HFD-fed Ldlr-/- mice resulted in hypocholesterolemic effects and significantly attenuated the progression of atherosclerosis. We found that L654 also reduced liver inflammatory and extracellular matrix gene expression, which may be related to inhibition of macrophage activation as demonstrated in vivo by lower major histocompatibility complex class II gene expression and confirmed in cell experiments. In addition, L654 and other bacterial glycine lipids in feces, liver, and serum were markedly reduced alongside changes in Bacteroidetes relative abundance in HFD-fed mice. Finally, we tested the bioactivities of L654 and related lipid 567 in chow-fed Apoe-/- mice, which displayed much higher fecal glycine lipids relative to HFD-fed Ldlr-/- mice. Administration of L654 or lipid 567 for 7 weeks to these mice reduced the liver injury marker alanine aminotransferase, but other effects seen in Ldlr-/- were not observed. Therefore, we conclude that conditions in which gut microbiome-derived glycine lipids are lost, such as HFD, may exacerbate the development of atherosclerosis and liver injury, whereas correction of such depletion may protect from these disorders.


Subject(s)
Atherosclerosis , Gastrointestinal Microbiome , Animals , Atherosclerosis/genetics , Bacteria , Bacteroidetes , Diet, High-Fat/adverse effects , Glycine/pharmacology , Liver , Mice , Mice, Inbred C57BL , Mice, Knockout , Serine
7.
Int J Mol Sci ; 23(5)2022 Feb 25.
Article in English | MEDLINE | ID: mdl-35269698

ABSTRACT

In addition to being involved in protein biosynthesis and metabolism, the amino acid glycine is the most important inhibitory neurotransmitter in caudal regions of the brain. These functions require a tight regulation of glycine concentration not only in the synaptic cleft, but also in various intracellular and extracellular compartments. This is achieved not only by confining the synthesis and degradation of glycine predominantly to the mitochondria, but also by the action of high-affinity large-capacity glycine transporters that mediate the transport of glycine across the membranes of presynaptic terminals or glial cells surrounding the synapses. Although most cells at glycine-dependent synapses express more than one transporter with high affinity for glycine, their synergistic functional interaction is only poorly understood. In this review, we summarize our current knowledge of the two high-affinity transporters for glycine, the sodium-dependent glycine transporters 1 (GlyT1; SLC6A9) and 2 (GlyT2; SLC6A5) and the alanine-serine-cysteine-1 transporter (Asc-1; SLC7A10).


Subject(s)
Glycine Plasma Membrane Transport Proteins , Synapses , Brain/metabolism , Glycine/pharmacology , Glycine Plasma Membrane Transport Proteins/metabolism , Neuroglia/metabolism , Synapses/metabolism
8.
Neuroimage ; 251: 119004, 2022 05 01.
Article in English | MEDLINE | ID: mdl-35176492

ABSTRACT

Although a substantial number of studies suggests some clinical benefit concerning negative symptoms in schizophrenia through the modulation of NMDA-receptor function, none of these approaches achieved clinical approval. Given the large body of evidence concerning glutamatergic dysfunction in a subgroup of patients, biomarkers to identify those with a relevant clinical benefit through glutamatergic modulation are urgently needed. A similar reduction of the early auditory evoked gamma-band response (aeGBR) as found in schizophrenia patients can be observed in healthy subjects following the application of an NMDA-receptor antagonist in the ketamine-model, which addresses the excitation / inhibition (E/I) imbalance of the disease. Moreover, this oscillatory change can be related to the emergence of negative symptoms. Accordingly, this study investigated whether glycine-related increases of the aeGBR, through NMDA-receptor co-agonism, accompany an improvement concerning negative symptoms in the ketamine-model. The impact of subanesthetic ketamine doses and the pretreatment with glycine was examined in twenty-four healthy male participants while performing a cognitively demanding aeGBR paradigm with 64-channel electroencephalography. Negative Symptoms were assessed through the PANSS. S-Ketamine alone caused a reduction of the aeGBR amplitude associated with more pronounced negative symptoms compared to placebo. Pretreatment with glycine attenuated both, the ketamine-induced alterations of the aeGBR amplitude and the increased PANSS negative scores in glycine-responders, classified based on relative aeGBR increase. Thus, we propose that the aeGBR represents a possible biomarker for negative symptoms in schizophrenia related to insufficient glutamatergic neurotransmission. This would allow to identify patients with negative symptoms, who might benefit from glutamatergic treatment.


Subject(s)
Glycine , Ketamine , Schizophrenia , Evoked Potentials, Auditory/drug effects , Evoked Potentials, Auditory/physiology , Glycine/pharmacology , Humans , Ketamine/adverse effects , Ketamine/pharmacology , Male , Receptors, N-Methyl-D-Aspartate , Schizophrenia/drug therapy
9.
Sci Rep ; 12(1): 2518, 2022 02 15.
Article in English | MEDLINE | ID: mdl-35169256

ABSTRACT

Stacking multiple genes into cotton crop to cop up multiple biotic stresses such as insects and weeds is a promising tool to save crop from losses. Transgenic cotton variety, VH-289, with double Bt and cp4EPSPS genes under the control of 35S promoter was used for the expression analyses and biosafety studies. The transgenic cotton plants were screened through PCR amplification of fragments, 1.7 kb for Cry1Ac, 582 bp for Cry2A and 250 bp for cp4EPSPS; which confirmed the presence of all genes transformed in transgenic cotton. The Cry1Ac + Cry2A and cp4EPSPS proteins were quantified through ELISA in transgenic cotton plants. The Glyphosate assay performed by spraying 1900 mL per acre of glyphosate Roundup further confirmed complete survival of transgenic cotton plants as compared to the non-transgenic cotton plants and all weeds. Similarly, insect infestation data determined that almost 99% insect mortality was observed in controlled field grown transgenic cotton plants as compared to the non-transgenic control plants. Evaluation of effect of temperature and soil nutrients availability on transgene expression in cotton plants was done at two different cotton growing regions, Multan and Lahore, Pakistan and results suggested that despite of higher temperature in Multan field, an increased level of Cry and cp4EPSPS proteins was recorded due to higher soil organic matter availability compared to Lahore field. Before commercialization of any transgenic variety its biosafety study is mandatory so, a 90 days biosafety study of the transgenic cotton plants with 40% transgenic cottonseeds in standard diet showed no harmful effect on wister rat model when studied for liver function, renal function and serum electrolyte.


Subject(s)
Glycine/analogs & derivatives , Gossypium/drug effects , Gossypium/genetics , Herbicide Resistance/genetics , Herbicides/pharmacology , Lepidoptera , Plant Weeds/drug effects , Animals , Diet/methods , Endotoxins/genetics , Endotoxins/metabolism , Glycine/pharmacology , Gossypium/metabolism , Hemolysin Proteins/genetics , Hemolysin Proteins/metabolism , Larva , Liver Function Tests , Male , Models, Animal , Pakistan , Plant Leaves/genetics , Plant Leaves/metabolism , Plants, Genetically Modified/drug effects , Plants, Genetically Modified/metabolism , Rats , Rats, Wistar , Risk Assessment , Seeds/genetics , Seeds/metabolism , Transgenes
10.
Molecules ; 27(3)2022 Feb 08.
Article in English | MEDLINE | ID: mdl-35164384

ABSTRACT

Anemia is a major complication of chronic renal failure. To treat this anemia, prolylhydroxylase domain enzyme (PHD) inhibitors as well as erythropoiesis-stimulating agents (ESAs) have been used. Although PHD inhibitors rapidly stimulate erythropoietin (Epo) production, the precise sites of Epo production following the administration of these drugs have not been identified. We developed a novel method for the detection of the Epo protein that employs deglycosylation-coupled Western blotting. With protein deglycosylation, tissue Epo contents can be quantified over an extremely wide range. Using this method, we examined the effects of the PHD inhibitor, Roxadustat (ROX), and severe hypoxia on Epo production in various tissues in rats. We observed that ROX increased Epo mRNA expression in both the kidneys and liver. However, Epo protein was detected in the kidneys but not in the liver. Epo protein was also detected in the salivary glands, spleen, epididymis and ovaries. However, both PHD inhibitors (ROX) and severe hypoxia increased the Epo protein abundance only in the kidneys. These data show that, while Epo is produced in many tissues, PHD inhibitors as well as severe hypoxia regulate Epo production only in the kidneys.


Subject(s)
Erythropoietin/metabolism , Glycine/analogs & derivatives , Isoquinolines/pharmacology , Prolyl-Hydroxylase Inhibitors/pharmacology , Protein Biosynthesis/drug effects , Animals , Erythropoietin/analysis , Erythropoietin/genetics , Female , Glycine/pharmacology , Hypoxia/genetics , Hypoxia/metabolism , Kidney/drug effects , Kidney/metabolism , Male , Rats , Rats, Sprague-Dawley , Up-Regulation/drug effects
11.
Org Lett ; 24(6): 1388-1393, 2022 02 18.
Article in English | MEDLINE | ID: mdl-35138108

ABSTRACT

A hybrid trans-AT PKS/NRPS gene cluster htm was identified with defined boundaries for hangtaimycin biosynthesis in Streptomyces spectabilis CPCC200148. Deoxyhangtaimycin, a new derivative of hangtaimycin, was identified from the htm11 gene knockout mutant. In vitro biochemical assays demonstrated that the cytochrome P450 monooxygenase Htm11 was responsible for the stereoselective hydroxylation of deoxyhangtaimycin to hangtaimycin. More importantly, deoxyhangtaimycin showed activity against influenza A virus at the micromolar level, highlighting its potential as an antiviral lead compound.


Subject(s)
Antiviral Agents/metabolism , Cytochrome P-450 Enzyme System/metabolism , Glycine/metabolism , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Biocatalysis , Glycine/chemistry , Glycine/pharmacology , Hydroxylation , Influenza A virus/drug effects , Microbial Sensitivity Tests , Molecular Structure
12.
Biochem Pharmacol ; 197: 114939, 2022 03.
Article in English | MEDLINE | ID: mdl-35114188

ABSTRACT

Kidney injury often causes anemia due to a lack of production of the erythroid growth factor erythropoietin (EPO) in the kidneys. Roxadustat is one of the first oral medicines inducing EPO production in patients with renal anemia by activating hypoxia-inducible factors (HIFs), which are activators of EPO gene expression. In this study, to develop prodrugs of roxadustat with improved permeability through cell membrane, we investigated the effects of 8 types of esterification on the pharmacokinetics and bioactivity of roxadustat using Hep3B hepatoma cells that HIF-dependently produce EPO. Mass spectrometry of cells incubated with the esterified roxadustat derivatives revealed that the designed compounds were deesterified after being taken up by cells and showed low cytotoxicity compared to the original compound. Esterification prolonged the effective duration of roxadustat with respect to EPO gene induction and HIF activation in cells transiently exposed to the compounds. In the kidneys and livers of mice, both of which are unique sites of EPO production, a majority of the methyl-esterified roxadustat was deesterified within 6 h after drug administration. The deesterified roxadustat derivative was continuously detectable in plasma and urine for at least 48 h after administration, while the administered compound became undetectable 24 h after administration. Additionally, we confirmed that methyl-esterified roxadustat activated erythropoiesis in mice by inducing Epo mRNA expression exclusively in renal interstitial cells, which have intrinsic EPO-producing potential. These data suggest that esterification could lead to the development of roxadustat prodrugs with improvements in cell membrane permeability, effective duration and cytotoxicity.


Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , Cell Survival/drug effects , Glycine/analogs & derivatives , Hypoxia-Inducible Factor-Proline Dioxygenases/metabolism , Intracellular Membranes/metabolism , Isoquinolines/metabolism , Isoquinolines/pharmacology , Animals , Basic Helix-Loop-Helix Transcription Factors/agonists , Cell Survival/physiology , Dose-Response Relationship, Drug , Esterification/drug effects , Esterification/physiology , Glycine/metabolism , Glycine/pharmacology , Humans , Intracellular Membranes/drug effects , Male , Mice , Mice, Inbred C57BL , Mice, Inbred ICR , Time Factors , Treatment Outcome , Tumor Cells, Cultured
13.
J Vis Exp ; (179)2022 01 10.
Article in English | MEDLINE | ID: mdl-35068479

ABSTRACT

Glyphosate-based products (GBP) are the most common broad-spectrum herbicides worldwide. The target of glyphosate is the enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) in the shikimate pathway, which is virtually universal in plants. The inhibition of the enzyme stops the production of three essential amino acids: phenylalanine, tyrosine, and tryptophan. EPSPS is also present in fungi and prokaryotes, such as archaea and bacteria; thus, the use of GBP may have an impact on the microbiome composition of soils, plants, herbivores, and secondary consumers. This article aims to present general guidelines to assess the effect of GBP on microbiomes from field experiments to bioinformatics analyses and provide a few testable hypotheses. Two field experiments are presented to test the GBP on non-target organisms. First, plant-associated microbes from 10 replicated control and GBP treatment plots simulating no-till cropping are sampled and analyzed. In the second experiment, samples from experimental plots fertilized by either poultry manure containing glyphosate residues or non-treated control manure were obtained. Bioinformatics analysis of EPSPS protein sequences is utilized to determine the potential sensitivity of microbes to glyphosate. The first step in estimating the effect of GBP on microbiomes is to determine their potential sensitivity to the target enzyme (EPSPS). Microbial sequences can be obtained either from public repositories or by means of PCR amplification. However, in the majority of field studies, microbiome composition has been determined based on universal DNA markers such as the 16S rRNA and the internal transcribed spacer (ITS). In these cases, sensitivity to glyphosate can only be estimated through a probabilistic analysis of EPSPS sequences using closely related species. The quantification of the potential sensitivity of organisms to glyphosate, based on the EPSPS enzyme, provides a robust approach for further experiments to study target and non-target resistant mechanisms.


Subject(s)
Herbicides , Microbiota , 3-Phosphoshikimate 1-Carboxyvinyltransferase/genetics , 3-Phosphoshikimate 1-Carboxyvinyltransferase/metabolism , Glycine/analogs & derivatives , Glycine/pharmacology , Herbicide Resistance/genetics , Herbicides/pharmacology , RNA, Ribosomal, 16S
14.
Cytogenet Genome Res ; 161(12): 578-584, 2021.
Article in English | MEDLINE | ID: mdl-35021177

ABSTRACT

In agriculture, various chemicals are used to control the weeds. Out of which, glyphosate is an important herbicide invariably used in the cultivation of glyphosate-resistant crops to control weeds. Overuse of glyphosate results in the evolution of glyphosate-resistant weeds. Evolution of glyphosate resistance (GR) in Amaranthus palmeri (AP) is a serious concern in the USA. Investigation of the mechanism of GR in AP identified different resistance mechanisms of which 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene amplification is predominant. Molecular analysis of GR AP identified the presence of a 5- to >160-fold increase in copies of the EPSPS gene than in a glyphosate-susceptible (GS) population. This increased copy number of the EPSPS gene increased the genome size ranging from 3.5 to 11.8%, depending on the copy number compared to the genome size of GS AP. FISH analysis using a 399-kb EPSPS cassette derived from bacterial artificial chromosomes (BACs) as probes identified that amplified EPSPS copies in GR AP exist in extrachromosomal circular DNA (eccDNA) in addition to the native copy in the chromosome. The EPSPS gene-containing eccDNA having a size of ∼400 kb is termed EPSPS-eccDNA and showed somatic mosacism in size and copy number. EPSPS-eccDNA has a genetic mechanism to tether randomly to mitotic or meiotic chromosomes during cell division or gamete formation and is inherited to daughter cells or progeny generating copy number variation. These eccDNAs are stable genetic elements that can replicate and exist independently. The genomic characterization of the EPSPS locus, along with the flanking regions, identified the presence of a complex array of repeats and mobile genetic elements. The cytogenomics approach in understanding the biology of EPSPS-eccDNA sheds light on various characteristics of EPSPS-eccDNA that favor GR in AP.


Subject(s)
Amaranthus/drug effects , Amaranthus/genetics , Cytogenetics , Genome, Plant/genetics , Glycine/analogs & derivatives , Herbicide Resistance/genetics , 3-Phosphoshikimate 1-Carboxyvinyltransferase/genetics , Amaranthus/cytology , DNA Copy Number Variations/genetics , Glycine/pharmacology , Plant Weeds/drug effects , Plant Weeds/genetics
15.
PLoS One ; 17(1): e0262494, 2022.
Article in English | MEDLINE | ID: mdl-35020774

ABSTRACT

Avena fatua and A. ludoviciana (commonly known as wild oats) are the most problematic winter grass species in fallows and winter crops in the northeast region of Australia. A series of experiments were conducted to evaluate the performance of glyphosate and alternative post-emergence herbicides on A. fatua and A. ludoviciana. This study reports the world's first glyphosate-resistant (GR) biotypes of A. fatua and A. ludoviciana. The glyphosate dose required to kill 50% of the plants (LD50) and to reduce 50% of the biomass (GR50) for the GR biotype of A. fatua was 556 g a.e./ha and 351 g a.e./ha, respectively. These values for A. ludoviciana were 848 g a.e./ha and 289 g a.e./ha. Regardless of the growth stage (3-4 or 6-7 leaf stages), clethodim (120 g a.i./ha), haloxyfop (78 g a.i./ha), pinoxaden (20 g a.i./ha), and propaquizafop (30 g a.i./ha) were the best alternative herbicide options for the control of A. fatua and A. ludoviciana. The efficacy of butroxydim (45 g a.i./ha), clodinafop (120 g a.i./ha), imazamox + imazapyr (36 g a.i./ha), and paraquat (600 g a.i./ha) reduced at the advanced growth stage. Glufosinate (750 g a.i./ha), flamprop (225 g a.i./ha), and pyroxsulam + halauxifen (20 g a.i./ha) did not provide effective control of Avena species. This study identified alternative herbicide options to manage GR biotypes of A. fatua and A. ludoviciana.


Subject(s)
Avena/growth & development , Crops, Agricultural/growth & development , Glycine/analogs & derivatives , Herbicide Resistance , Herbicides/pharmacology , Avena/classification , Avena/drug effects , Crops, Agricultural/drug effects , Glycine/pharmacology
16.
Int J Mol Sci ; 23(1)2021 Dec 25.
Article in English | MEDLINE | ID: mdl-35008638

ABSTRACT

Rigosertib is multi-kinase inhibitor that could represent an interesting therapeutic option for non-resectable patients with cholangiocarcinoma, a very aggressive hepatic cancer with limited effective treatments. The Western blotting technique was used to evaluate alterations in the expression of proteins involved in the regulation of the cell cycle of cholangiocarcinoma EGI-1 cells. Our results show an increase in EMI1 and Cyclin B protein levels after Rigosertib treatment. Moreover, the phosphorylation of CDK1 is significantly reduced by Rigosertib, while PLK1 expression increased after 24 h of treatment and decreased after 48 h. Finally, we evaluated the role of p53. Its levels increase after Rig treatment, and, as shown in the cell viability experiment with the p53 inhibitor Pifithrin, its activity is necessary for the effects of Rigosertib against the cell viability of EGI-1 cells. In conclusion, we hypothesized the mechanism of the action of Rigosertib against cholangiocarcinoma EGI-1 cells, highlighting the importance of proteins involved in the regulation of cell cycles. The CDK1-Cyclin B complex and p53 play an important role, explaining the Block in the G2/M phase of the cell cycle and the effect on cell viability.


Subject(s)
Cell Division/drug effects , Cholangiocarcinoma/drug therapy , G2 Phase/drug effects , Glycine/analogs & derivatives , Sulfones/pharmacology , CDC2 Protein Kinase/metabolism , Cell Cycle Proteins/metabolism , Cholangiocarcinoma/metabolism , Cyclin B/metabolism , Glycine/pharmacology , Humans , Phosphorylation/drug effects , Protein Kinase Inhibitors/pharmacology , Proto-Oncogene Proteins/metabolism , Signal Transduction/drug effects , Tumor Cells, Cultured , Tumor Suppressor Protein p53/metabolism
17.
ACS Chem Neurosci ; 13(2): 229-244, 2022 01 19.
Article in English | MEDLINE | ID: mdl-34990110

ABSTRACT

The activation of N-methyl-d-aspartate receptor (NMDAR) is triggered by the closure of bilobed (D1 and D2) clamshell-like clefts upon binding glycine (Gly) and glutamate. There is evidence that cholinergic compounds modulate NMDAR-mediated currents via direct receptor-ligand interactions; however, molecular bases are unknown. Here, we first propose a mechanistic structure-based explanation for the observed ACh-induced submaximal potentiation of NMDA-elicited currents in striatal neurons by predicting competitive inhibition with Gly. Then, the model was validated, in principle, by confirming that the coapplication of Gly and ACh significantly reduces these neuronal currents. Finally, we delineate the interplay of ACh with the NMDAR by a combination of computational strategies. Crystallographic ACh-bound complexes were studied, revealing a similar ACh binding environment on the GluN1 subunit of the NMDAR. We illustrate how ACh can occupy X-ray monomeric open, dimeric "semiopen" cleft conformations obtained by molecular dynamics and a full-active cryo-EM NMDAR structure, explaining the suboptimal NMDAR electrophysiological activity under the "Venus Flytrap model". At an evolutionary biology level, the binding mode of ACh coincides with that of the homologous ornithine-bound periplasmic LAO binding protein complex. Our computed results indicate an analogous mechanism of action, inasmuch as ACh may stabilize the GluN1 subunit "semiclosed" conformations by inducing direct and indirect D1-to-D2 interdomain bonds. Additionally, an alternative binding site was detected, shared by the known NMDAR allosteric modulators. Experimental and computed results strongly suggest that ACh acts as a Gly-competitive, submaximal potentiating agent of the NMDAR, possibly constituting a novel chemotype for multitarget-directed drug development, e.g., to treat Alzheimer's, and it may lead to a new understanding of glutamatergic neurotransmission.


Subject(s)
Acetylcholine , Receptors, N-Methyl-D-Aspartate , Glycine/pharmacology , N-Methylaspartate , Neurons
18.
Cancer Treat Rev ; 103: 102334, 2022 Feb.
Article in English | MEDLINE | ID: mdl-34974243

ABSTRACT

Isocitrate dehydrogenase 1 (IDH1) has been investigated as a promising therapeutic target in select cancers with a mutated version of the enzyme (mtIDH1). With only one phase III trial published to date and two indications approved for routine clinical use by the FDA, we reviewed the entire clinical trial portfolio to broadly understand mtIDH1 inhibitor activity in patients. We queried PubMed.gov and ClinicalTrials.gov to identify published and ongoing clinical trials related to IDH1 and cancer. Progression-free survival (PFS), overall survival (OS), 2-hydroxyglutarate levels, and adverse events were summarized. To date, ten clinical trials investigating mtIDH1 inhibitors among patients with diverse malignancies (cholangiocarcinoma, acute myeloid leukemia, chondrosarcoma, glioma) have been published. Almost every trial (80%) has investigated ivosidenib. In multiple phase I trials, ivosidenib treatment resulted in promising radiographic and biochemical responses with improved survival outcomes (relative to historic data) among patients with both solid and hematologic mtIDH1 malignancies. Among patients enrolled in a phase III trial with advanced cholangiocarcinoma, ivosidenib resulted in a PFS rate of 32% at 6 months, as compared to 0% with placebo. There was a 5.2 month increase in OS with ivosidenib relative to placebo, after considering crossover. The treatment-specific grade ≥3 adverse event rate of ivosidenib was 2%-26% among all patients, and was just 3.6% among 284 patients who had a solid tumor across four trials. Although <1% of malignancies harbor IDH1 mutations, small molecule mtIDH1 inhibitors, namely ivosidenib, appear to be biologically active and well tolerated in patients with solid and hematologic mtIDH1 malignancies.


Subject(s)
Antineoplastic Agents/therapeutic use , Enzyme Inhibitors/therapeutic use , Glycine/analogs & derivatives , Isocitrate Dehydrogenase/antagonists & inhibitors , Neoplasms/drug therapy , Pyridines/therapeutic use , Aniline Compounds/adverse effects , Aniline Compounds/pharmacology , Aniline Compounds/therapeutic use , Antineoplastic Agents/adverse effects , Antineoplastic Agents/pharmacology , Benzimidazoles/adverse effects , Benzimidazoles/pharmacology , Benzimidazoles/therapeutic use , Clinical Trials as Topic , Enzyme Inhibitors/adverse effects , Enzyme Inhibitors/pharmacology , Glycine/adverse effects , Glycine/pharmacology , Glycine/therapeutic use , Humans , Isocitrate Dehydrogenase/genetics , Mutation , Neoplasms/mortality , Pyridines/adverse effects , Pyridines/pharmacology
19.
Environ Microbiol Rep ; 14(1): 70-84, 2022 02.
Article in English | MEDLINE | ID: mdl-34786867

ABSTRACT

Roundup® is the brand name for herbicide solutions containing glyphosate, which specifically inhibits the 5-enolpyruvyl-shikimate-3-phosphate (EPSP) synthase of the shikimate pathway. The inhibition of the EPSP synthase causes plant death because EPSP is required for biosynthesis of aromatic amino acids. Glyphosate also inhibits the growth of archaea, bacteria, Apicomplexa, algae and fungi possessing an EPSP synthase. Here, we have characterized two glyphosate-resistant bacteria from a Roundup solution. Taxonomic classification revealed that the isolates 1CH1 and 2CH1 are Burkholderia anthina and Burkholderia cenocepacia strains respectively. Both isolates cannot utilize glyphosate as a source of phosphorus and synthesize glyphosate-sensitive EPSP synthase variants. Burkholderia. anthina 1CH1 and B. cenocepacia 2CH1 tolerate high levels of glyphosate because the herbicide is not taken up by the bacteria. Previously, it has been observed that the exposure of soil bacteria to herbicides like glyphosate promotes the development of antibiotic resistances. Antibiotic sensitivity testing revealed that the only the B. cenocepacia 2CH1 isolate showed increased resistance to a variety of antibiotics. Thus, the adaptation of B. anthina 1CH1 and B. cenocepacia 2CH1 to glyphosate did not generally increase the antibiotic resistance of both bacteria. However, our study confirms the genomic adaptability of bacteria belonging to the genus Burkholderia.


Subject(s)
3-Phosphoshikimate 1-Carboxyvinyltransferase , Burkholderia cenocepacia , 3-Phosphoshikimate 1-Carboxyvinyltransferase/chemistry , 3-Phosphoshikimate 1-Carboxyvinyltransferase/metabolism , Burkholderia , Burkholderia cenocepacia/genetics , Burkholderia cenocepacia/metabolism , Glycine/analogs & derivatives , Glycine/chemistry , Glycine/pharmacology
20.
Amino Acids ; 54(3): 385-398, 2022 Mar.
Article in English | MEDLINE | ID: mdl-33839961

ABSTRACT

Glycine is an amino acid with a diverse array of health benefits regarding metabolism, immunity, and development. The aim of this study was to test the hypothesis that glycine supplementation alters the intestinal microbial composition and improves the intestinal mucosal immunity of weaned piglets. One hundred and twenty-eight weaned piglets divided into 4 groups were fed with a corn- and soybean meal-based diet supplemented with 0 (control), 0.5, 1, or 2% glycine for 7 days. The intestinal microbiota and tissue samples from the control and the 2% glycine-supplemented piglets were collected for determination of the composition of microbial community and the intestinal mucosal barrier function. Piglets fed with diet containing 2% glycine, instead of 0.5% or 1% glycine, presented elevated average daily gain and feed conversion ratio, as compared with the control. 2% glycine enhanced the abundance of mucins in the jejunum and ileum and mRNA level of porcine ß-defensin (pBD) 2 and pBD-3, as well as the protein level of secretory immunoglobulin A (sIgA) in the jejunum. The mRNA expression of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6, and the protein level of phosphorylated p38 mitogen-activated protein kinase (MAPK), signal transducer and activator of transcription 3 (STAT3), nuclear factor (NF)-κB p65, and claudin-2 in the jejunum were lower in the 2% glycine group than that in the control. In addition, an elevated ratio of CD4+/CD8+ T lymphocytes was observed in the jejunum of piglets receiving diet supplemented with 2% glycine. The colon content of piglets fed with 2% glycine exhibited a reduction in abundance of pathogenic bacteria (Escherichia-Shigella, Clostridium, and Burkholderiales) and an increase in short-chain fatty acid-producing bacteria (Blautia, Lachnospiraceae, Anaerostipes, and Prevotella) in comparison with the control. We conclude that dietary supplementation with 2% glycine improves the intestinal immunological barrier function and the microbial composition, therefore, contributing to the growth performance of weaned piglets.


Subject(s)
Glycine , Immunity, Mucosal , Animals , Dietary Supplements , Glycine/metabolism , Glycine/pharmacology , Intestinal Mucosa/metabolism , Intestines , Swine , Weaning
SELECTION OF CITATIONS
SEARCH DETAIL