Transgenic Nematodes Research Articles

Loss of DNA glycosylases improves health and cognitive function in a C. elegans model of human tauopathy.

Alzheimer's disease (AD) is a neurodegenerative disorder representing a major burden on families and society. Some of the main pathological hallmarks of AD are the accumulation of amyloid plaques (Aβ) and tau neurofibrillary tangles. However, it is still unclear how Aβ and tau aggregates promote specific phenotypic outcomes and lead to excessive oxidative DNA damage, neuronal cell death and eventually to loss of memory. Here we utilized a Caenorhabditis elegans (C. elegans)model of human tauopathy to investigate the role of DNA glycosylases in disease development and progression. Transgenic nematodes expressing a pro-aggregate form of tau displayed altered mitochondrial content, decreased lifespan, and cognitive dysfunction. Genetic ablation of either of the two DNA glycosylases found in C. elegans, NTH-1 and UNG-1, improved mitochondrial function, lifespan, and memory impairment. NTH-1 depletion resulted in a dramatic increase of differentially expressed genes, which was not apparent in UNG-1 deficient nematodes. Our findings clearly show that in addition to its enzymatic activity, NTH-1 has non-canonical functions highlighting its modulation as a potential therapeutic intervention to tackle tau-mediated pathology.

Neurotoxicity of tetrabromobisphenol-A-bis(2,3-dibromopropyl ether) through the GABAergic and serotonergic neurotransmission in Caenorhabditis elegans

Tetrabromobisphenol-A-bis(2,3-dibromopropyl ether) (TBBPA-BDBPE), a novel additive brominated flame retardant, is being developed for use in polyolefin and copolymers. Despite its emerging application, the neurotoxicity and mechanisms of action of TBBPA-BDBPE remain unexplored. Caenorhabditis elegans was utilized as the model organism to study the neurotoxic effects of TBBPA-BDBPE across environmental concentrations ranging from 0 to 100 μg/L. This investigation focused on various toxicological endpoints such as locomotive behavior, neuronal injury, neurotransmitter transmission, and the regulation of nervous system-related gene expression. Acute exposure to TBBPA-BDBPE at concentrations of 10–100 μg/L significantly impaired nematode movement, indicating potential neurotoxicity. In transgenic nematodes, this exposure also caused damage to γ-aminobutyric acid (GABAergic) and serotonergic neurons, along with notable changes in the levels of GABAergic and serotonergic neurotransmitters. Further molecular studies indicated alterations in neurotransmission-related genes (cat-4, mod-1, unc-25, and unc-47). Molecular docking analysis confirmed the binding affinity of TBBPA-BDBPE to key neurotransmission proteins—CAT-4, MOD-1, UNC-25, and UNC-47. These findings demonstrate that TBBPA-BDBPE exerts neurotoxic effects by impacting GABAergic and serotonergic neurotransmission in nematodes. This study provides new insights into the potential environmental risks of TBBPA-BDBPE.

2,4,6-trinitrotoluene causes mitochondrial toxicity in Caenorhabditis elegans by affecting electron transport

As a typical energetic compound widely used in military activities, 2,4,6-trinitrotoluene (TNT) has attracted great attention in recent years due to its heavy pollution and wide distribution in and around the training facilities, firing ranges, and demolition sites. However, the subcellular targets and the underlying toxic mechanism of TNT remain largely unknown. In this study, we explored the toxic effects of TNT biological reduction on the mitochondrial function and homeostasis in Caenorhabditis elegans (C. elegans). With short-term exposure of L4 larvae, 10–1000 ng/mL TNT reduced mitochondrial membrane potential and adenosine triphosphate (ATP) content, which was associated with decreased expression of specific mitochondrial complex involving gas-1 and mev-1 genes. Using fluorescence-labeled transgenic nematodes, we found that fluorescence expression of sod-3 (muls84) and gst-4 (dvls19) was increased, suggesting that TNT disrupted the mitochondrial antioxidant defense system. Furthermore, 10 ng/mL TNT exposure increased the expression of the autophagy-related gene pink-1 and activated mitochondrial unfolded protein response (mt UPR), which was indicated by the increased expression of mitochondrial stress activated transcription factor atfs-1, ubiquitin-like protein ubl-5, and homeobox protein dve-1. Our findings demonstrated that TNT biological reduction caused mitochondrial dysfunction and the development of mt UPR protective stress responses, and provided a basis for determining the potential risks of energetic compounds to living organisms.

Integrated UHPLC-Q-Orbitrap-MS/MS Method and Network Pharmacology for Exploring the Active Components and Potential Mechanisms of Neuroprotective Effect of the n-Butanol Part of Mume Flos

Background: Mume Flos is a common Qi-regulating medication used in traditional Chinese medicine, modern study has discovered that Mume Flos and its components have neuroprotective properties. However, the effective components and molecular mechanisms of Mume Flos are still unclear. Methods: In this study, the current study chose Caenorhabditis elegans as a model to evaluate the anti-Alzheimer disease (AD) effect of the active parts of Mume Flos using life span, paralysis rate, oxidative stress capacity, and heat stress capacity as indicators. Then, UHPLC-Q-Orbitrap-MS/MS and network pharmacology combined with GEO database chip were used for joint analysis to predict the active components and potential molecular mechanisms, which were validated using C. elegans models. Results: The results showed that the n-butanol part of Mume Flos slowed down nematode paralysis and possessed anti-AD activity, and we tentatively identified a total of 45 constituents, mainly including flavonoids and phenylpropanoids as well as a small number of amino acids, and determined that 39 active components and 130 targets are related to neuroprotection. Further analysis identified the main active ingredients such as rutin, neochlorogenic acid, and chlorogenic acid, all of which could reduce Aβ deposition in nematodes and slow down the rate of paralysis in transgenic nematodes. Conclusion: This study revealed the neuroprotective effects of MFB and identified its active components as rutin, neochlorogenic acid, and chlorogenic acid. They may play a role in reducing Aβ deposition by regulating the targets of GAPDH, TNF, IL6, INS, and TP53. This study provides a basis for further in-depth research on the mechanism of MFB to improve neurological diseases.

Human wild-type and D76N β2-microglobulin variants are significant proteotoxic and metabolic stressors for transgenic C. elegans.

β2-microglobulin (β2-m) is a plasma protein derived from physiological shedding of the class I major histocompatibility complex (MHCI), causing human systemic amyloidosis either due to persistently high concentrations of the wild-type (WT) protein in hemodialyzed patients, or in presence of mutations, such as D76N β2-m, which favor protein deposition in the adulthood, despite normal plasma levels. Here we describe a new transgenic Caenorhabditis elegans (C. elegans) strain expressing human WT β2-m at high concentrations, mimicking the condition that underlies dialysis-related amyloidosis (DRA) and we compare it to a previously established strain expressing the highly amyloidogenic D76N β2-m at lower concentrations. Both strains exhibit behavioral defects, the severity of which correlates with β2-m levels rather than with the presence of mutations, being more pronounced in WT β2-m worms. β2-m expression also has a deep impact on the nematodes' proteomic and metabolic profiles. Most significantly affected processes include protein degradation and stress response, amino acids metabolism, and bioenergetics. Molecular alterations are more pronounced in worms expressing WT β2-m at high concentration compared to D76N β2-m worms. Altogether, these data show that β2-m is a proteotoxic protein invivo also in its wild-type form, and that concentration plays a key role in modulating pathogenicity. Our transgenic nematodes recapitulate the distinctive features subtending DRA compared to hereditary β2-m amyloidosis (high levels of non-mutated β2-m vs. normal levels of variant β2-m) and provide important clues on the molecular bases of these human diseases.

Comparative oxidative damages induced by silver nanoparticles with different sizes and coatings in Caenorhabditis elegans.

Silver nanoparticles (AgNPs) are widely used in many commercial and medical products. Serious concerns are paid on their adverse potentials to the environment and human health. In this study, toxic effects and oxidative stress induced by AgNPs with different sizes and coatings (20nm AgNPs, 20nm polyvinylpyrrolidone (PVP) -AgNPs and 50nm AgNPs) in Caenorhabditis elegans (C. elegans) were investigated. The toxic effects including the shortened lifespan and decreased frequency of head thrashes and body bends of C. elegans were induced in a dose-dependent manner by AgNPs. The reactive oxygen species (ROS) production and the oxidative stress-related indicators including malondialdehyde (MDA) and glutathione (GSH) in nematodes were changed after exposure to three kinds of AgNPs. These effects were the most obvious in a 20nm PVP-AgNPs exposure group. AgNPs could also induce the expression of genes related to oxidative stress in nematodes. In addition, the up-regulation of mtl-1 and mtl-2 in nematodes might reduce the oxidative damage caused by AgNPs, by using transgenic strains CF2222 and CL2120 nematodes. Metallothionein (MT), an antioxidant, could relieve the oxidative damage caused by AgNPs. These results suggested that 20nm PVP-AgNPs with a smaller particle size and better dispersion have stronger toxic effects and the oxidative damage to nematodes. Mtl-1 and mtl-2 might be involved in alleviating the oxidative damage caused by AgNPs. Our findings provide clues for the safety evaluation and mechanism information of metal nanoparticles.

Soybean Cyst Nematode Management Is Improved by Combining Native and Transgenic Resistance.

Field trials were conducted to assess the benefit of combining a transgenic soybean cyst nematode (SCN) resistance trait, Cry14Ab-1 expressed by the event GMB151, with the native resistance allele rhg1b from PI 88788. The GMB151 event and rhg1b were crossed into common genetic backgrounds and segregated out to create four genetically related lines within each background. The lines created contained both native and transgenic resistance (rhg1b + GMB151), only native resistance (rhg1b alone), only transgenic resistance (GMB151 alone), or neither resistance type (susceptible). The benefit of GMB151 and rhg1b for SCN management was evaluated by measuring SCN control and yield protection. Soybean cyst nematode control was assessed by counting the number of females and cysts on roots early in the season and measuring the change in SCN egg population density over the entire season. The GMB151 transgenic event and the native resistance allele rhg1b both reduced early season SCN reproduction and contributed to significantly higher soybean yield. Compared to susceptible lines, the rhg1b allele improved yield by 33%, while GMB151 improved yield by 13%. Combining the GMB151 event and rhg1b allele resulted in greater SCN control and yield improvement than either provided alone. The combination of GMB151 and rhg1b reduced season-long SCN reproduction by 50% and resulted in 44% greater yield than the susceptible lines. Soybean cyst nematode virulence to rhg1b continues to increase due to the continuous planting of PI 88788-derived resistant cultivars. Pyramiding GMB151 with rhg1b provides a new management option to improve SCN control and soybean yield.

Repurposing Antimicrobial Protegrin-1 as a Dual-Function Amyloid Inhibitor via Cross-seeding.

Amyloids and antimicrobial peptides have traditionally been recognized as distinct families with separate biological functions and targets. However, certain amyloids and antimicrobial peptides share structural and functional characteristics that contribute to the development of neurodegenerative diseases. Specifically, the aggregation of amyloid-β (Aβ) and microbial infections are interconnected pathological factors in Alzheimer's disease (AD). In this study, we propose and demonstrate a novel repurposing strategy for an antimicrobial peptide of protegrin-1 (PG-1), which exhibits the ability to simultaneously prevent Aβ aggregation and microbial infection both in vitro and in vivo. Through a comprehensive analysis using protein, cell, and worm assays, we uncover multiple functions of PG-1 against Aβ, including the following: (i) complete inhibition of Aβ aggregation at a low molar ratio of PG-1/Aβ = 0.25:1, (ii) disassembly of the preformed Aβ fibrils into amorphous aggregates, (iii) reduction of Aβ-induced cytotoxicity in SH-SY5Y cells and transgenic GMC101 nematodes, and (iv) preservation of original antimicrobial activity against P.A., E.coli., S.A., and S.E. strains in the presence of Aβ. Mechanistically, the dual anti-amyloid and anti-bacterial functions of PG-1 primarily arise from its strong binding to distinct Aβ seeds (KD = 1.24-1.90 μM) through conformationally similar β-sheet associations. This work introduces a promising strategy to repurpose antimicrobial peptides as amyloid inhibitors, effectively targeting multiple pathological pathways in AD.

Photoaged microplastics induce neurotoxicity associated with damage to serotonergic, glutamatergic, dopaminergic, and GABAergic neuronal systems in Caenorhabditis elegans

Microplastics (MPs) are ubiquitous environmental contaminants that cause neurotoxicity in various organisms. MPs are typically affected by light irradiation and undergo photoaging. However, the neurotoxic effects of photoaged polystyrene (P-PS) and its underlying mechanisms remain unclear. In this study, locomotion behaviors, neuronal development, neurotransmitter levels, and the expression of neurotransmission-related genes were investigated in Caenorhabditis elegans exposed to P-PS at environment-relevant concentrations (0.1–100 μg/L). The characterization results showed that photoaging accelerated the aging process and changed the physicochemical properties of the MPs. The toxicity results suggested that exposure to 1–100 μg/L P-PS caused more severe neurotoxicity than virgin polystyrene (V-PS) with endpoints of head thrashes, body bends, wavelength, and mean amplitude. Exposure to P-PS also altered the fluorescence intensity and neurodegeneration percentage of serotonergic, glutamatergic, dopaminergic, and aminobutyric acid (GABA) in transgenic nematodes. Similarly, significant reductions in the levels of these neurotransmitters were also observed. Based on Pearson's correlation, locomotion behaviors were negatively correlated with the neurotransmission of serotonin, glutamate, dopamine, and GABA. Further investigation suggested that the expression of neurotransmitter-related genes (e.g., tph-1, eat-4, and unc-46) was significantly altered in the nematodes. Collectively, the neurotoxic effects of P-PS were attributed to abnormal neurotransmission. This study highlights the potential toxicity of MPs photoaged under environmentally relevant conditions.

Ethyl caffeate attefnuates Aβ-induced toxicity in Caenorhabditis elegans AD models via the insulin/insulin-like growth factor-1 signaling pathway

The pathogenesis of Alzheimer's disease (AD), a multifactorial progressive neurodegenerative disease associated with aging, is unclear. Ethyl caffeate is a plant polyphenol that has been reported to have neuroprotective effects, but the mechanisms by which it acts are unclear. In this study, for the first time, we investigated the molecular mechanism of its anti-AD properties using the Caernorhabditis elegans model. The results of our experiments showed that ethyl caffeate delayed the paralysis symptoms of CL4176 to a different extent and reduced the exogenous 5-hydroxytryptophan-induced paralysis phenotype. Further studies revealed that ethyl caffeate lowered Aβ plaques and depressed the expression of Aβ monomers and oligomers, but did not influence the mRNA levels of Aβ. Moreover, it was able to bring paraquat-induced ROS levels down to near-standard conditions. Real-time quantitative PCR experiment showed a significant upregulation of the transcript abundance of daf-16, skn-1 and hsf-1, key factors associated with the insulin/insulin-like growth factor 1 (IGF-1) signaling pathway (IIS), and their downstream genes sod-3, gst-4 and hsp-16.2. It was further shown that ethyl caffeate activated the translocation of DAF-16 and SKN-1 from the cytoplasm to the nucleus and enhanced the expression of sod-3::GFP, gst-4::GFP and hsp-16.2::GFP in transgenic nematodes. This meant that the protection against Aβ toxicity by ethyl caffeate may be partly through the IIS signaling pathway. In addition, ethyl caffeate suppressed the aggregation of polyglutamine proteins in AM141, which indicated a potential protective effect against neurodegenerative diseases based on abnormal folding and aggregation of amyloid proteins. Taken together, ethyl caffeate is expected to develop as a potential drug for the management of AD.

Synthesis, anti-aging and mechanism of magnolol derivatives.

Magnolol (M), a hydroquinone containing an allyl side chain, is one of the major active components of Houpoea officinalis for antioxidation and anti-aging. To enhance the antioxidant activity of magnolol, the different sites of magnolol were structurally modified in this experiment, and a total of 12 magnolol derivatives were obtained. Based on the preliminary exploration of the anti-aging effect of magnolol derivatives in a Caenorhabditis elegans (C. elegans) model. Our results indicate that the active groups of magnolol exerting anti-aging effects were allyl groups and hydroxyl on the phenyl. Meanwhile, the anti-aging effect of the novel magnolol derivative M27 was found to be significantly superior to that of magnolol. To investigate the effect of M27 on senescence and the potential mechanism of action, we investigated the effect of M27 on senescence in C. elegans. In this study, we investigated the effect of M27 on C. elegans physiology by examining body length, body curvature and pharyngeal pumping frequency. The effect of M27 on stress resistance in C. elegans was explored by acute stress experiments. The mechanism of M27 anti-aging was investigated by measuring ROS content, DAF-16 nuclear translocation, sod-3 expression, and lifespan of transgenic nematodes. Our results indicate that M27 prolonged the lifespan of C. elegans. Meanwhile, M27 improved the healthy lifespan of C. elegans by improving pharyngeal pumping ability and reducing lipofuscin accumulation in C. elegans. M27 increased resistance to high temperature and oxidative stress in C. elegans by reducing ROS. M27 induced DAF-16 translocation from cytoplasm to nucleus in transgenic TJ356 nematodes and upregulated the expression of sod-3 (a gene downstream of DAF-16) in CF1553 nematodes. Furthermore, M27 did not extend the lifespan of daf-16, age-1, daf-2, and hsp-16.2 mutants. This work suggests that M27 may ameliorate aging and extend lifespan in C. elegans through the IIS pathway.

Polygonati Rhizoma Polysaccharide Prolongs Lifespan and Healthspan in Caenorhabditis elegans.

Polygonati Rhizoma is the dried rhizome of Polygonatum kingianum coll.et hemsl., Polygonatum sibiricum Red. or Polygonatum cyrtonema Hua, and has a long history of medication. Raw Polygonati Rhizoma (RPR) numbs the tongue and stings the throat, while prepared Polygonati Rhizoma (PPR) can remove the numbness of the tongue, and at the same time enhance its functions of invigorating the spleen, moistening the lungs and tonifying the kidneys. There are many active ingredients in Polygonati Rhizoma (PR), among which polysaccharide is one of the most important active ingredients. Therefore, we studied the effect of Polygonati Rhizoma polysaccharide (PRP) on the lifespan of Caenorhabditis elegans (C. elegans) and found that polysaccharide in PPR (PPRP) was more effective than Polysaccharide in RPR (RPRP) in prolonging the lifespan of C. elegans, reducing the accumulation of lipofuscin, and increasing the frequency of pharyngeal pumping and movement. The further mechanism study found that PRP can improve the anti-oxidative stress ability of C. elegans, reduce the accumulation of reactive oxygen species (ROS) in C. elegans, and improve the activity of antioxidant enzymes. The results of quantitative real-time PCR(q-PCR) experiments suggested that PRP may prolong the lifespan of C. elegans by down-regulating daf-2 and activating daf-16 and sod-3, and the transgenic nematode experiments were consistent with its results, so it was hypothesized that the mechanism of age delaying effect of PRP was related to daf-2, daf-16 and sod-3 of the insulin signaling pathway. In short, our research results provide a new idea for the application and development of PRP.

Biomonitoring of Indoor Air Fungal or Chemical Toxins with Caenorhabditis elegans nematodes.

Bad indoor air quality due to toxins and other impurities can have a negative impact on human well-being, working capacity and health. Therefore, reliable methods to monitor the health risks associated with exposure to hazardous indoor air agents are needed. Here, we have used transgenic Caenorhabditis elegans nematode strains carrying stress-responsive fluorescent reporters and evaluated their ability to sense fungal or chemical toxins, especially those that are present in moisture-damaged buildings. Liquid-based or airborne exposure of nematodes to mycotoxins, chemical agents or damaged building materials reproducibly resulted in time- and dose-dependent fluorescent responses, which could be quantitated by either microscopy or spectrometry. Thus, the C. elegans nematodes present an easy, ethically acceptable and comprehensive in vivo model system to monitor the response of multicellular organisms to indoor air toxicity.

Carboxyl-modified polystyrene microplastics induces neurotoxicity by affecting dopamine, glutamate, serotonin, and GABA neurotransmission in Caenorhabditis elegans

Microplastics (MPs) are ubiquitous in various environmental media and have potential toxicity. However, the neurotoxicity of carboxyl-modified polystyrene microplastics (PS-COOH) and their mechanisms remain unclear. In this study, Caenorhabditis elegans was used as a model to examine the neurotoxicity of polystyrene microplastic (PS) and PS-COOH concentrations ranging from 0.1 to 100 μg/L. Locomotion behavior, neuron development, neurotransmitter level, and neurotransmitter-related gene expression were selected as assessment endpoints. Exposure to low concentrations (1 μg/L) of PS-COOH caused more severe neurotoxicity than exposure to pristine PS. In transgenic nematodes, exposure to PS-COOH at 10–100 μg/L significantly increased the fluorescence intensity of dopaminergic, glutamatergic, serotonergic, and aminobutyric acid (GABA)ergic neurons compared to that of the control. Further studies showed that exposure to 100 μg/L PS-COOH can significantly affect the levels of glutamate, serotonin, dopamine, and GABA in nematodes. Likewise, in the present study, the expression of genes involved in neurotransmission was altered in worms. These results suggest that PS-COOH exerts neurotoxicity by affecting neurotransmission of dopamine, glutamate, serotonin, and GABA. This study provides new insights into the underlying mechanisms and potential risks associated with PS-COOH.

LC-MS/MS Insight into Vitamin C Restoration to Metabolic Disorder Evoked by Amyloid β in Caenorhabditis elegans CL2006.

The transitional expression and aggregation of amyloid β (Aβ) are the most important causative factors leading to the deterioration of Alzheimer’s disease (AD), a commonly occurring metabolic disease among older people. Antioxidant agents such as vitamin C (Vc) have shown potential effects against AD and aging. We applied an liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method and differential metabolites strategy to explore the metabolic disorders and Vc restoration in a human Aβ transgenic (Punc-54::Aβ1–42) nematode model CL2006. We combined the LC-MS/MS investigation with the KEGG and HMDB databases and the CFM-ID machine-learning model to identify and qualify the metabolites with important physiological roles. The differential metabolites responding to Aβ activation and Vc treatment were filtered out and submitted to enrichment analysis. The enrichment showed that Aβ mainly caused abnormal biosynthesis and metabolism pathways of phenylalanine, tyrosine and tryptophan biosynthesis, as well as arginine and proline metabolism. Vc reversed the abnormally changed metabolites tryptophan, anthranilate, indole and indole-3-acetaldehyde. Vc restoration affected the tryptophan metabolism and the biosynthesis of phenylalanine, tyrosine and tryptophan. Our findings provide supporting evidence for understanding the metabolic abnormalities in neurodegenerative diseases and the repairing effect of drug interventions.

P14-08 Antiobesity and antioxidant activity of cocoa beans and shells on wild and transgenic nematodes

P14-08 Antiobesity and antioxidant activity of cocoa beans and shells on wild and transgenic nematodes

Exendin-4 alleviates β-Amyloid peptide toxicity via DAF-16 in a Caenorhabditis elegans model of Alzheimer's disease.

Epidemiological analyses indicate that type 2 diabetes mellitus (T2DM) is a risk factor for Alzheimer's disease (AD). They share common pathophysiological mechanisms. Thus, it has been increasingly suggested that several anti-T2DM drugs may have therapeutic potential in AD. Exendin-4, as a glucagon-like peptide-1 (GLP-1) receptor agonist, is an approved drug used to treat T2DM. In this research, the neuroprotective effect of Exendin-4 was investigated for the first time using transgenic Caenorhabditis elegans. Our results demonstrated that Exendin-4 attenuated the amyloid-β (1-42) (Aβ1-42) toxicity via multiple mechanisms, such as depressing its expression on protein and mRNA and reducing Aβ (1-42) accumulation. Exendin-4 at 0.5 mg/ml had been shown to extend life by 34.39% in CL4176 and delay the onset of paralysis in CL4176 and CL2006 which were increased by 8.18 and 8.02%, respectively. With the treatment of Exendin-4, the nuclear translocation of DAF-16 in the transgenic nematode TJ356 was enhanced. Superoxide dismutase-3 (SOD-3), as a downstream target gene regulated by DAF-16, was upregulated on mRNA level and activity. The reactive oxygen species (ROS) level was decreased. In contrast, we observed that the ability of Exendin-4 to regulate SOD was decreased in CL4176 worms with the DAF-16 gene silenced. The activity of SOD and the mRNA level of sod-3 were downregulated by 30.45 and 43.13%, respectively. Taken together, Exendin-4 attenuated Aβ (1-42) toxicity in the C. elegans model of AD via decreasing the expression and the accumulation of Aβ (1-42). Exendin-4 exhibited the ability of antioxidant stress through DAF-16. With continuous research, Exendin-4 would become a potential therapeutic strategy for treating AD.

Anti-Alzheimer's disease active components screened out and identified from Hedyotis diffusa combining bioaffinity ultrafiltration LC-MS with acetylcholinesterase

Ethnopharmacological relevanceHedyotis diffusa is a traditional ethnomedicinal plant in local communities in northeastern Asia and used to treat inflammation, nervous breakdown, among others. In recent years, it has been applied in the treatment of Alzheimer's disease (AD), while the specific chemical components responsible for the activity remain need to be explored. Aim of the studyTo prepare, screen and identify the potential anti-AD active components from Hedyotis diffusa. Materials and methodsThe acetylcholinesterase (AChE) inhibitory activity of four different extracts of Hedyotis diffusa were initially assessed using a spectrophotometric Ellman's method. A more accurate LC-MS/MS screening method combining functional enzyme assay and affinity ultrafiltration (AU) screening assay was developed and applied for the screening of natural compound inhibitors of AChE from Hedyotis diffusa. The binding mode was further investigated between protein and ligands via molecular docking. Subsequently, CL4176, a transgenic nematode model for AD, was used for activity validation of one of these components. ResultsN-butanol extract of Hedyotis diffusa (NHD) appeared significant inhibitory activities on AChE, were chosen to delve deeper. Five bioactive components targeting AChE were screened out and identified using AU coupled to liquid chromatography-mass spectrometry. Molecular docking technique further confirmed the results of the screening assay. Finally, quercetin-3-O-sophoroside (QS) was confirmed as a potent anti-AD agent by in vivo experiments in C. elegans. ConclusionThis study explores a new idea for screening anti-AD active components from traditional medicine. The findings provide a molecular structure and bioactivity basis for future potential applications of Hedyotis diffusa in medical industries.

Neurotoxicity of Tris (1,3-dichloroisopropyl) phosphate in Caenorhabditis elegans

As a new type of flame retardant, Organic Phosphate Flame Retardant has been widely used worldwide. The purpose of our research is to determine the neurotoxicity of Tris (1,3-dichloroisopropyl) phosphate (TDCPP) to Caenorhabditis elegans and its mechanism. L1 larvae wild-type C. elegans were exposed to different concentrations of TDCPP, and the effects on motor behavior (head thrashes, body bends, pumping times, chemotaxis index), ROS levels, and p38MAPK signaling pathway-related gene expression levels were measured. Three transgenic nematode strains, BZ555, DA1240, and EG1285, were also used to study the effects of TDCPP on nematode dopamine neurons, glutamate neurons, and GABA neurons. The results showed that TDCPP can inhibit the head thrashes and body bends of the nematode, reduce dopamine production, increase the level of ROS in the body, and affect the expression of genes related to the p38MAPK signaling pathway. We next employed ROS production and motor behavior as toxicity assessment endpoints to determine the involvement of p38 MAPK signaling in the regulation of response to TDCPP. The results showed that the nematodes with low expression of pmk-1 were less sensitive to the TDCPP. It was suggested that TDCPP had neurotoxicity and regulated neurotoxicity to C. elegans by activating the p38-MAPK signaling pathway. The research in this article provides important information for revealing the environmental health risks of organophosphorus flame retardants and their toxic mechanism of action.

Design of Gallic Acid-Glutamine Conjugate and Chemical Implications for Its Potency Against Alzheimer's Amyloid-β Fibrillogenesis.

Epigallocatechin-3-gallate (EGCG) has been widely recognized as a potent inhibitor of Alzheimer's amyloid-β (Aβ) fibrillogenesis. We found that gallic acid (GA) has superior inhibitory effects over EGCG at the same mass concentrations and assumed the pivotal role of the carboxyl group in GA. Therefore, we designed five GA-derivatives to investigate the significance of carboxyl groups in modulating Aβ fibrillogenesis, including carboxyl-amidated GA (GA-NH2), GA-glutamic acid conjugate (GA-E), and GA-E derivatives with amidated either of the two carboxyl groups (GA-Q and GA-E-NH2) or with two amidated-carboxyl groups (GA-Q-NH2). Intriguingly, only GA-Q shows significantly stronger potency than GA and extends the life span of the AD transgenic nematode by over 30%. Thermodynamic studies reveal that GA-Q has a strong binding affinity for Aβ42 with two binding sites, one stronger (site 1, Ka1 = 3.1 × 106 M-1) and the other weaker (site 2, Ka2 = 0.8 × 106 M-1). In site 1, hydrogen bonding, electrostatic interactions, and hydrophobic interactions all have contributions, while in site 2, only hydrogen bonding and electrostatic interactions work. The two sites are confirmed by molecular simulations, and the computations specified the key residues. GA-Q has strong binding to Asp23, Gly33, Gly38, Ala30, Ile31, and Leu34 via hydrogen bonding and electrostatic interactions, while it interacts with Phe19, Ala21 Gly25, and Asn27 via hydrophobic interactions. Consequently, GA-Q destroys Asp23-Lys28 salt bridges and restricts β-sheet/bridge structures. The thermodynamic and molecular insight into the GA-Q functions on inhibiting Aβ fibrillogenesis would pave a new way to the design of potent molecules against Alzheimer's amyloid.