Alzheimer's disease (AD) implicates neuronal loss, plaque and neurofibrillary tangle formation, and disturbed neuronal Ca(2+) homeostasis, which leads to severe dementia, memory loss, as well as thinking and behavioral perturbations that could ultimately lead to death. Calcium dysregulation and low acetylcholine levels are two main mechanisms implicated in Alzheimer's disease progression. Simultaneous inhibition of calcium oscillations (store overload-induced Ca(2+) release [SOICR]) and acetylcholinesterase (AChE) by a single molecule may bring a new breath of hope for AD treatment. Here, we described some dantrolene derivatives as dual inhibitors of the ryanodine receptor and AChE. Two series of acylhydrazone/sulfonylhydrazone derivatives with aromaticgroup were designed and synthesized. In this study, the target compounds were evaluated for their ability to inhibit SOICR and AChE in vitro, using dantrolene and donepezil as positive controls. Compound 22a exhibited excellent and balanced inhibitory potency against SOICR (inhibition (%) = 90.1, IC(50) = 0.162 microM) and AChE (inhibition (%) = 93.5, IC(50) = 0.372 microM). Docking simulations showed that several preferred compounds could bind to the active sites of both the proteins, further validating the rationality of the design strategy. Potential therapeutic effects in AD were evaluated using the Barnes maze and Morris water maze tests, which demonstrated that compound 22a significantly improved memory and cognitive behavior in AD model mice. Moreover, it was also found that compound 22a could enhance synaptic strength by measuring hippocampal long-term potentiation (LTP) in brain slices. These results suggested that the introduction of a sulfonyl-hydrazone scaffold and aromatic substitution to dantrolene derivatives provided a useful template for the development of potential chemical entities against AD.
Title: Diketopyrrolopyrrole-based fluorescent probe for visualizing over-expressed carboxylesterase in fever via ratiometric imaging Zhang B, Qin S, Wang N, Lu X, Jiao J, Zhang J, Zhao W Ref: Talanta, 266:124971, 2023 : PubMed
Fever is the result of inflammation and the innate self-defense response of organisms, can cause abnormal changes in the activity of many enzymes in organisms, including the important carboxylesterase (CE). Monitoring the activity changes of CE in vivo during a fever will help to understand heat-related pathological mechanisms. In this paper, we designed diketopyrrolopyrrole-based ratiometric fluorescent probes DPP-FBC-P and DPP-FBO-P containing alkyl chain and diethylene glycol monomethyl ether chain respective for detection of CE. Both probes could realized fast response to CE and displayed good selectivity and high sensitivity. Compared with DPP-FBO-P, DPP-FBC-P had better biocompatibility, larger signal to noise ratio (225-fold vs 125-fold) and lower detection limit (1.6 x 10(-5) U/mL vs 4.2 x 10(-5) U/mL). Moreover, the probe DPP-FBC-P had been successfully applied to image the endogenous CE in HepG2 cells and solid tumors, and also visualized the over expressed CE in fever cells. Most importantly, the changes of CE level in the liver of fever mice model induced by LPS were monitored with the assistance of DPP-FBC-Pvia dual channel ratio imaging for the first time. In addition, fluorescence color signal in solution was captured by smart phone, and the linear relationship between RGB ratio (G/R) and CE concentration was established. This work will provide a potential approach for investigating the physiological and pathological processes of heat related diseases.
Multi-targeted directed ligands (MTDLs) are emerging as promising Alzheimer's disease (AD) therapeutic possibilities. Coumarin is a multifunctional backbone with extensive bioactivity that has been utilized to develop innovative anti-neurodegenerative properties and is a desirable starting point for the construction of MTDLs. Herein, we explored and synthesized a series of novel coumarin derivatives and assessed their inhibitory effects on cholinesterase (AChE, BuChE), GSK-3beta, and BACE1. Among these compounds, compound 30 displayed the multifunctional profile of targeting the AChE (IC(50) = 1.313 +/- 0.099 microM) with a good selectivity over BuChE (SI = 24.623), GSK-3beta (19.30% inhibition at 20 microM), BACE1 (IC(50) = 1.227 +/- 0.112 microM), along with moderate HepG2 cytotoxicity, SH-SY5Y cytotoxicity, low HL-7702 cytotoxicity, as well as good blood-brain barrier (BBB) permeability. Kinetic and docking studies indicated that compound 30 was a competitive AChE inhibitor. Furthermore, acute toxicity experiments revealed that it was non-toxic at a dosage of 1000 mg/kg. The ADME prediction results indicate that 30 has acceptable physicochemical properties. Collectively, these findings demonstrated that compound 30 would be a potential multifunctional candidate for AD therapy.
Alzheimer's disease (AD) is characterized by progressive cognitive impairment and mental behavior. The combination inhibition of two essential AD targets, acetylcholinesterase (AChE) and glycogen synthase kinase-3beta (GSK-3beta), might be a breakthrough in the discovery of therapeutic success. Herein, 17 beta-carboline-1,2,3-triazole hybrids were designed, synthesized, and evaluated for their AChE and GSK-3beta inhibitory potential. The results indicated that compound 21 has the most potent inhibition against eeAChE (IC(50) = 0.20 +/- 0.02 microM), hAChE (IC(50) = 0.34 +/- 0.01 microM) and GSK-3beta (IC(50) = 1.14 +/- 0.05 microM) among these compounds. In addition, it inhibited hAChE in a mixed type manner and could occupy the binding pocket forming diverse interactions with the target of AChE and GSK-3beta. Moreover, compound 21 showed low cytotoxicity against SH-SY5Y and HepG2 cell lines and good BBB permeability. Compound 21 also attenuated the tau hyperphosphorylation in the Tau (P301L) 293T cell model. The ADME projection exhibited that compound 21 has acceptable physicochemical characteristics. This study provides new leads for the assessment of AChE and GSK-3beta dual inhibition as a promising strategy for AD treatment.
Title: Co, N co-doped porous carbon-based nanozyme as an oxidase mimic for fluorescence and colorimetric biosensing of butyrylcholinesterase activity Sun W, Wang N, Zhou X, Sheng Y, Su X Ref: Mikrochim Acta, 189:363, 2022 : PubMed
A Co, N co-doped porous carbon-based nanozyme (Co-N-C nanozyme) has been fabricated. Taking advantages of the excellent oxidase catalytic activity and significant stability of Co-N-C nanozyme, we propose a fluorescence and colorimetric system based on Co-N-C nanozyme and red-emitting carbon quantum dots (RCDs) for butyrylcholinesterase (BChE) sensing. As the chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) was catalyzed and oxidized by Co-N-C nanozyme, the generated oxTMB had a new absorption peak at 652 nm, which resulted in the significant quenching of the fluorescence of the carbon quantum dots at 610 nm. Under the catalysis of BChE, thiocholine was generated from the hydrolysis of S-butyrylthiocholine iodide (BTCh), and the as-generated thiocholine effectively inhibited the oxidation of TMB catalyzed by Co-N-C nanozyme, leading to a decrease of the absorption of oxTMB at 652 nm and effective fluorescence recovery of RCDs. By measuring the absorbance of produced oxTMB at 652 nm and the fluorescence of RCDs at 610 nm, the fluorescence and colorimetric system both exhibited an outstanding linear response to the activity of BChE in the range 0.5 to 40 U L(-1), with a detection limit of 0.16 U L(-1) and 0.21 U L(-1), respectively. Furthermore, this established dual-channel biosensing strategy has been successfully applied to the determination of BChE in human serum samples. The present work has effectively expanded the development and application of nanozyme in biosensing.
Title: Constructing bifunctional metal-organic framework based nanozymes with fluorescence and oxidase activity for the dual-channel detection of butyrylcholinesterase Wang N, Shi J, Liu Y, Sun W, Su X Ref: Anal Chim Acta, 1205:339717, 2022 : PubMed
Herein, we reported a novel strategy for the fabrication of bifunctional metal-organic framework based nanozymes (oxidized UiO-66-NH(2)@Ce), which displayed excellent oxidase mimic activity as well as fluorescence property. The bifunctional oxidized UiO-66-NH(2)@Ce possess excellent oxidase activity due to oxidase-like active Ce(4+)/Ce(3+) sites, which makes the nanozymes have strong positive charge, resulting in a stronger affinity for the negatively charged chromogenic substrate 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). Utilizing the bifunctional oxidized UiO-66-NH(2)@Ce, a sensitive fluorometric and colorimetric dual-channel detection strategy for butyrylcholinesterase (BChE) was fabricated for the first time. The oxidized UiO-66-NH(2)@Ce could catalyze the oxidation of colorless ABTS to green oxABTS, which in turn quench the fluorescence of oxidized UiO-66-NH(2)@Ce. Butyrylcholinesterase (BChE) can catalyze the hydrolysis of S-butyrylthiocholine iodide (BTCh) to produce thiocholine, which could prevent the oxidation of ABTS, resulting in the fluorescence of oxidized UiO-66-NH(2)@Ce recovered. Both the colorimetric and fluorometric dual-channel sensing platform displayed a sensitive response to BChE, and the limits of detection (LOD) for BChE could achieve as low as 0.056 and 0.050U/L, respectively. The dual-output assay for BChE detection displayed excellent application prospects.
CDK2/9 are members of the CDKs family, which play key roles in the occurrence and development of many cancers by regulating cell cycle and transcriptional prolongation, respectively. To further optimize and discuss the structure-activity relationships (SARs), a series of tacrine-based compounds were designed and synthesized from the compound ZLWT-37, which was studied by our group previously but no detailed SARs study was conducted on CDK2/9. Among this series, compounds ZLMT-12 (35) exhibited the most potent antiproliferative activity (GI(50) = 0.006 microM for HCT116) and superior CDK2/9 inhibitory properties (CDK2: IC(50) = 0.011 microM, CDK9: IC(50) = 0.002 microM). Meanwhile, ZLMT-12 showed a weak inhibitory effect on acetylcholinesterase (AChE, IC(50) = 19.023 microM) and butyrylcholinesterase (BuChE, IC(50) = 2.768 microM). In addition, ZLMT-12 can suppress colony formation and migration in HCT116 cells, as well as induce the apoptosis and arrest the cell cycle in the S phase and G2/M phase. In vivo investigations revealed that ZLMT-12 inhibits tumor growth in the HCT116 xenograft tumor model at a low dose of 10 mg/kg without causing hepatotoxicity. The acute toxicity test showed low toxicity with a median lethal dosage (LD(50)) of 104.417 mg/kg. These findings showed that ZLMT-12 might be used as a drug candidate by targeting CDK2/9.
Title: Mutation in BrGGL7 gene encoding a GDSL esterase / lipase causes male sterility in Chinese cabbage (Brassica rapa L. ssp. pekinensis) Zhao Y, Huang S, Zou J, Dong S, Wang N, Feng H Ref: Theor Appl Genet, _135:3323, 2022 : PubMed
MutMap and KASP analyses revealed that the BrGGL7 gene is responsible for the male-sterile trait of ftms1 in Chinese cabbage, with functional verification in Arabidopsis. The application of a male-sterile line is an ideal approach of hybrid seed production in Chinese cabbage. In this study, we obtained a male-sterile mutant (ftms1) from the double haploid line 'FT' using ethyl methane sulfonate (EMS) mutagenesis. The mutant was completely sterile due to abnormal enlargement and vacuolization of the tapetum cells. A single recessive nuclear gene was found to control male sterility in the mutant, while MutMap and KASP analyses identified BraA05g022470.3C (BrGGL7), which encodes a GDSL esterase / lipase, as the candidate mutant gene. A single nucleotide substitution from C to T occurred within the domain of BrGGL7 in ftms1, resulting in premature translation termination in the fourth exon. Meanwhile, qRT-PCR analysis indicated that BrGGL7 was prominently expressed in the anothers, and expression was greater in the wild-type 'FT' than ftms1. Genetic complementation of the orthologous Arabidopsis ggl7 mutant further confirmed the role of BrGGL7 in pollen development. These findings suggest that BrGGL7 plays a fundamental role in pollen formation, providing important insight into the molecular mechanisms underlying male sterility in Chinese cabbage.
The natural product harmine, a representative beta-carboline alkaloid from the seeds of Peganum harmala L. (Zygophyllaceae), possesses a broad spectrum of biological activities. In this study, a novel series of harmine derivatives containing N-benzylpiperidine moiety were identified for the treatment of Alzheimer's disease (AD). The results showed that all the derivatives possessed significant anti-acetylcholinesterase (AChE) activity and good selectivity over butyrylcholinesterase (BChE). In particular, compound ZLWH-23 exhibited potent anti-AChE activity (IC(50) = 0.27 microM) and selective BChE inhibition (IC(50) = 20.82 microM), as well as acceptable glycogen synthase kinase-3 (GSK-3beta) inhibition (IC(50) = 6.78 microM). Molecular docking studies and molecular dynamics simulations indicated that ZLWH-23 could form stable interaction with AChE and GSK-3beta. Gratifyingly, ZLWH-23 exhibited good selectivity for GSK-3beta over multi-kinases and very low cytotoxicity towards SH-SY5Y, HEK-293T, HL-7702, and HepG2 cell lines. Importantly, ZLWH-23 displayed efficient reduction against tau hyperphosphorylation on Ser-396 site in Tau (P301L) 293T cell model. Collectively, harmine-based derivatives could be considered as possible drug leads for the development of AD therapies.
Title: Early enteral nutrition combined with PSS-based nursing in the treatment of organophosphorus pesticide poisoning Sun Y, Yang Y, Zhang Z, Li Y, Hu Y, Wang N Ref: Am J Transl Res, 13:9315, 2021 : PubMed
OBJECTIVE: To investigate the administration of early enteral nutrition combined with poisoning severity score (PSS)-based nursing in the treatment of organophosphorus pesticide poisoning (OPP). METHODS: A total of 99 OPP patients treated in our hospital between June 2019 and June 2020 were enrolled in this study and were divided into the conventional group (n=46, early enteral nutrition support + routine care) and the combined group (n=53, PSS-based nursing + early enteral nutrition support + routine care). The nutritional status indicators, the hemoglobin (Hb) and blood glucose levels, the Glasgow coma scale (GCS) scores, and the complications were compared between the two groups. RESULTS: The total protein (TP), albumin (ALB), and prealbumin (PAB) levels were reduced in the conventional group after the intervention (P<0.05) but were significantly lower than they were in the combined group (P<0.05). The Hb and blood glucose levels were decreased in the conventional group after the intervention (P<0.05) and were significantly higher than they were in the combined group (P<0.05). The GCS scores increased significantly as the treatment progressed (P<0.05), and the GCS scores in the combined group were significantly higher than the GCS scores in the conventional group at 3 and 5 days after the treatment (P<0.05). The time to the recovery of 60% cholinesterase (CHE) activity, the durations of the mechanical ventilation, the lengths of the hospital stays, and the hospital costs in the combined group were significantly lower than they were in the conventional group (P<0.05). The complication rate in the combined group (9.43%) was significantly lower than the complication rate in the conventional group (32.61%) (P<0.05). CONCLUSION: Early enteral nutrition combined with PSS-based nursing can effectively control the blood glucose, improve the nutritional disorders, promote recovery, and reduce complications in OPP patients.
Alzheimer's disease (AD) is a prevalent neurodegenerative disorder that has multiple causes. Therefore, multiple-target-directed ligands (MTDLs), which act on multiple targets, have been developed as a novel strategy for AD therapy. In this study, novel drug candidates were designed and synthesized by the covalent linkings of tacrine, a previously used anti-AD acetylcholinesterase (AChE) inhibitor, and dipicolylamine, an beta-amyloid (Abeta) aggregation inhibitor. Most tacrine-dipicolylamine dimers potently inhibited AChE and Abeta(1-42) aggregation in vitro, and 13a exhibited nanomolar level inhibition. Molecular docking analysis suggested that 13a could interact with the catalytic active sites and the peripheral anion site of AChE, and bind to Abeta(1-42) pentamers. Moreover, 13a effectively attenuated Abeta(1-42) oligomers-induced cognitive dysfunction in mice by activating the cAMP-response element binding protein/brain-derived neurotrophic factor signaling pathway, decreasing tau phosphorylation, preventing synaptic toxicity, and inhibiting neuroinflammation. The safety profile of 13a in mice was demonstrated by acute toxicity experiments. All these results suggested that novel tacrine-dipicolylamine dimers, especially 13a, have multi-target neuroprotective and cognitive-enhancing potentials, and therefore might be developed as MTDLs to combat AD.
Title: Magnetic COFs as satisfied support for lipase immobilization and recovery to effectively achieve the production of biodiesel by great maintenance of enzyme activity Zhou ZW, Cai CX, Xing X, Li J, Hu ZE, Xie ZB, Wang N, Yu XQ Ref: Biotechnol Biofuels, 14:156, 2021 : PubMed
BACKGROUND: Production of biodiesel from renewable sources such as inedible vegetable oils by enzymatic catalysis has been a hotspot but remains a challenge on the efficient use of an enzyme. COFs (Covalent Organic Frameworks) with large surface area and porosity can be applied as ideal support to avoid aggregation of lipase and methanol. However, the naturally low density limits its application. In this work, we reported a facile synthesis of core-shell magnetic COF composite (Fe(3)O(4)@COF-OMe) to immobilize RML (Rhizomucor miehei lipase), to achieve its utilization in biodiesel production. RESULT: This strategy gives extrinsic magnetic property, and the magnetic COFs is much heavier and could disperse in water medium well, facilitating the attachment with the enzyme. The resultant biocomposite exhibited an excellent capacity of RML due to its high surface area and fast response to the external magnetic field, as well as good chemical stability. The core-shell magnetic COF-OMe structure not only achieved highly efficient immobilization and recovery processes but also maintained the activity of lipase to a great extent. RML@Fe(3)O(4)@COF-OMe performed well in practical applications, while free lipase did not. The biocomposite successfully achieved the production of biodiesel from Jatropha curcas Oil with a yield of about 70% in the optimized conditions. CONCLUSION: Magnetic COFs (Fe(3)O(4)@COF-OMe) for RML immobilization greatly improved catalytic performance in template reaction and biodiesel preparation. The magneticity makes it easily recovered and separated from the system. This first successful attempt of COFs-based immobilized enzyme broadened the prospect of biodiesel production by COFs with some inspiration.
Generating chromosome-level, haplotype-resolved assemblies of heterozygous genomes remains challenging. To address this, we developed gamete binning, a method based on single-cell sequencing of haploid gametes enabling separation of the whole-genome sequencing reads into haplotype-specific reads sets. After assembling the reads of each haplotype, the contigs are scaffolded to chromosome level using a genetic map derived from the gametes. We assemble the two genomes of a diploid apricot tree based on whole-genome sequencing of 445 individual pollen grains. The two haplotype assemblies (N50: 25.5 and 25.8 Mb) feature a haplotyping precision of greater than 99% and are accurately scaffolded to chromosome-level.
Amanita neoovoidea (genus Amanita Pers.) poisoning leads to acute renal failure. Here, we present seven case reports of acute renal failure with acute hepatic failure due to ingestion of A. neoovoidea. Clinical manifestations included gastrointestinal symptoms 1-72 h after ingestion; elevation of renal parameters and blood uric acid, blood urea nitrogen, and creatinine levels; a few abnormal hepatic parameters, primarily albumin decrease and alanine aminotransferase increase; and elevation of zymogram parameters such as cholinesterase and lactate dehydrogenase. To determine whether the hepatic/renal lesions were caused by amanitins, we analyzed the blood and urine samples of patients and specimens of poisonous mushrooms. Morphological and molecular biological analyses indicated that the mushroom was A. neoovoidea. However, no amatoxins and phallotoxins were detected in its basidiomata.
Title: Enhancing secretion of polyethylene terephthalate hydrolase PETase in Bacillus subtilis WB600 mediated by the SP(amy) signal peptide Wang N, Guan F, Lv X, Han D, Zhang Y, Wu N, Xia X, Tian J Ref: Lett Appl Microbiol, :, 2020 : PubMed
The polyethylene terephthalate hydrolase (PETase) has been proved to have a high activity to degrade polyethylene terephthalate (PET), but few studies have been carried on its secretion in Bacillus subtilis. In this study, the coding gene of PETase, which was isolated from the Ideonella sakaiensis, was synthesized and expressed in B. subtilis. Then, we evaluated the ability of five Bacillus signal peptides to enhance PETase secretion by B. subtilis. The results indicated that the SP(amy) -induced secretion of PETase was the highest, and its activity against p-Nitrophenyl palmitate was about fourfold that of the natural signal peptide SP(PETase) . The weak promoter P43 provided sufficient time for translation and folding of PETase, resulting in increased extracellular expression. Use of P43 and SP(amy) in combination yielded the greatest bis-(2-hydroxyethyl) terephthalate degradation and PET-film etching activity due to maximized secretion of PETase by B. subtilis. Our findings will facilitate biodegradation of PET plastic.
Our recently successful identification of benzoic acid-based DPP-4 inhibitors spurs the further quest for in-depth structure-activity relationships (SAR) study in S2' site DPP-4. Thus novel benzamide fragments were designed to target the S2' site to compromise lipophilicity and improve oral activity. Exploring SAR by introduction of a variety of amide and halogen on benzene ring led to identification of several compounds, exerting moderated to excellent DPP-4 activities, in which 4'-chlorine substituted methyl amide 17g showed most potent DPP-4 activity with the IC50 value of 1.6nM. Its activity was superior to reference alogliptin. Docking study ideally verified and interpreted the obtained SAR of designed compounds. As a continuation, DPP-8/9 assays revealed the designed compounds exhibited good selectivity over DPP-8 and DPP-9. Subsequent cell-based test indicated compound 17g displayed low toxicity toward the LO2 cell line up to 100muM. In vivo evaluation showed compound 17g robustly improved the glucose tolerance in normal mice. Importantly, 17g exhibited reasonable pharmacokinetic (PK) profiles for oral delivery. Overall, compound 17g has the potential to a safe and efficacious DPP-4 inhibitor for T2DM treatment.
Dipeptidyl Peptidase-IV (DPP-4) is a validated therapeutic target for type 2 diabetes. Aiming to interact with both residues Try629 and Lys554 in S2' site, a series of novel uracil derivatives 1a-l and 2a-i incorporating benzoic acid moieties at the N3 position were designed and evaluated for their DPP-4 inhibitory activity. Structure-activity relationships (SAR) study led to the identification of the optimal compound 2b as a potent and selective DPP-4 inhibitor (IC50=1.7nM). Docking study revealed the additional salt bridge formed between the carboxylic acid and primary amine of Lys554 has a key role in the enhancement of the activity. Furthermore, compound 2b exhibited no cytotoxicity in human hepatocyte LO2 cells up to 50muM. Subsequent in vivo evaluations revealed that the ester of 2b robustly improves the glucose tolerance in normal mice. The overall results have shown that compound 2b has the potential to a safe and efficacious treatment for T2DM.
Six new dihydroisocoumarins, aspergimarins A-F (1-6), were discovered together with five known analogs (7-11) from a monoculture of the sponge-derived fungus Aspergillus sp. NBUF87. The structures of these compounds were elucidated through comprehensive spectroscopic methods, and absolute configurations were assigned after X-ray crystallography, use of the modified Mosher's method, and comparison of electronic circular dichroism (ECD) data with literature values for previously reported analogs. Compounds 1-11 were evaluated in a variety of bioassays, and at 100 muM, both 1 and 5 showed significant inhibitory effects on the lateral root growth of Arabidopsis thaliana Columbia-0 (Col-0). Moreover, at 100 muM, 5 also possessed notable inhibition against the primary root growth of Col-0. Meanwhile, 1-11 were all found to be inactive in vitro against acetylcholinesterase (AChE) (IC50 > 100 muM), four different types of human-derived cancer cell lines (IC50 > 50 muM), as well as methicillin-resistant Staphylococcus aureus and Escherichia coli (MIC > 50 mug/mL), and Plasmodium falciparum W2 (EC50 > 100 mug/mL), in phenotypic tests.
Title: Targeted acetylcholinesterase-responsive drug carriers with long duration of drug action and reduced hepatotoxicity Lin Y, Wang Y, Lv J, Wang N, Wang J, Li M Ref: Int J Nanomedicine, 14:5817, 2019 : PubMed
Purpose: Acetylcholinesterase (AChE) plays a critical role in the transmission of nerve impulse at the cholinergic synapses. Design and synthesis of AChE inhibitors that increase the cholinergic transmission by blocking the degradation of acetylcholine can serve as a strategy for the treatment of AChE-associated disease. Herein, an operational targeted drug delivery platform based on AChE-responsive system has been presented by combining the unique properties of enzyme-controlled mesoporous silica nanoparticles (MSN) with clinical-used AChE inhibitor. Methods: Functionalized MSNs were synthesized by liquid phase method and characterized by using different analytical methods. The biocompatibility and cytotoxicity of MSNs were determined by hemolysis experiment and MTT assay, respectively. Comparison of AChE activity between drug-loading system and inhibitor was developed with kits and by ELISA method. The efficacy of drug-loaded nanocarriers was investigated in a mouse model. Results: Compared with AChE inhibitor itself, the inhibition efficiency of this drug delivery system was strongly dependent on the concentration of AChE. Only AChE with high concentration could cause the opening of pores in the MSN, leading to the controlled release of AChE inhibitor in disease condition. Critically, the drug delivery system can not only exhibit long duration of drug action on AChE inhibition but also reduce the hepatotoxicity in vivo. Conclusion: In summary, AChE-responsive drug release systems have been far less explored. Our results would shed lights on the design of enzyme controlled-release multifunctional system for enzyme-associated disease treatment.
beta-Amyloid (Abeta) is regarded as an important pathogenic target for Alzheimer's disease (AD), the most prevalent neurodegenerative disease. Abeta can assemble into oligomers and fibrils, and produce neurotoxicity. Therefore, Abeta aggregation inhibitors may have anti-AD therapeutic efficacies. It was found, here, that the marine-derived alkaloid, fascaplysin, inhibits Abeta fibrillization in vitro. Moreover, the new analogue, 9-methylfascaplysin, was designed and synthesized from 5-methyltryptamine. Interestingly, 9-methylfascaplysin is a more potent inhibitor of Abeta fibril formation than fascaplysin. Incubation of 9-methylfascaplysin with Abeta directly reduced Abeta oligomer formation. Molecular dynamics simulations revealed that 9-methylfascaplysin might interact with negatively charged residues of Abeta42 with polar binding energy. Hydrogen bonds and pi(-)pi interactions between the key amino acid residues of Abeta42 and 9-methylfascaplysin were also suggested. Most importantly, compared with the typical Abeta oligomer, Abeta modified by nanomolar 9-methylfascaplysin produced less neuronal toxicity in SH-SY5Y cells. 9-Methylfascaplysin appears to be one of the most potent marine-derived compounds that produces anti-Abeta neuroprotective effects. Given previous reports that fascaplysin inhibits acetylcholinesterase and induces P-glycoprotein, the current study results suggest that fascaplysin derivatives can be developed as novel anti-AD drugs that possibly act via inhibition of Abeta aggregation along with other target mechanisms.
Middle East respiratory syndrome coronavirus (MERS-CoV) emerged into the human population in 2012 and has caused substantial morbidity and mortality. Potently neutralizing antibodies targeting the receptor-binding domain (RBD) on MERS-CoV spike (S) protein have been characterized, but much less is known about antibodies targeting non-RBD epitopes. Here, we report the structural and functional characterization of G2, a neutralizing antibody targeting the MERS-CoV S1 N-terminal domain (S1-NTD). Structures of G2 alone and in complex with the MERS-CoV S1-NTD define a site of vulnerability comprising two loops, each of which contain a residue mutated in G2-escape variants. Cell-surface binding studies and in vitro competition experiments demonstrate that G2 strongly disrupts the attachment of MERS-CoV S to its receptor, dipeptidyl peptidase-4 (DPP4), with the inhibition requiring the native trimeric S conformation. These results advance our understanding of antibody-mediated neutralization of coronaviruses and should facilitate the development of immunotherapeutics and vaccines against MERS-CoV.
Title: Recent Advances in Multi-target Anti-Alzheimer Disease Compounds (2013 Up to the Present) Wang N, Qiu P, Cui W, Yan X, Zhang B, He S Ref: Curr Med Chem, 26:5684, 2019 : PubMed
Since the last century, when scientists proposed the lock-and-key model, the discovery of drugs has focused on the development of drugs acting on single target. However, single-target drug therapies are not effective to complex diseases with multi-factorial pathogenesis. Moreover, the combination of single-target drugs readily causes drug resistance and side effects. In recent years, multi-target drugs have increasingly been represented among FDA-approved drugs. Alzheimer's Disease (AD) is a complex and multi-factorial disease for which the precise molecular mechanisms are still not fully understood. In recent years, rational multi-target drug design methods, which combine the pharmacophores of multiple drugs, have been increasingly applied in the development of anti-AD drugs. In this review, we give a brief description of the pathogenesis of AD and provide detailed discussions about the recent development of chemical structures of anti-AD agents (2013 up to present) that have multiple targets, such as amyloid-beta peptide, Tau protein, cholinesterases, monoamine oxidase, beta-site amyloid-precursor protein-cleaving enzyme 1, free radicals, metal ions (Fe2+, Cu2+, Zn2+) and so on. In this paper, we also added some novel targets or possible pathogenesis which have been reported in recent years for AD therapy. We hope that these findings may provide new perspectives for the pharmacological treatment of AD.
BACKGROUND: Hereditary spastic paraplegias (HSP) is a heterogeneous group of rare neurodegenerative disorders affecting the corticospinal tracts. To date, more than 78 HSP loci have been mapped to cause HSP. However, both the clinical and mutational spectrum of Chinese patients with HSP remained unclear. In this study, we aim to perform a comprehensive analysis of clinical phenotypes and genetic distributions in a large cohort of Chinese HSP patients, and to elucidate the primary pathogenesis in this population. METHODS: We firstly performed next-generation sequencing targeting 149 genes correlated with HSP in 99 index cases of our cohort. Multiplex ligation-dependent probe amplification testing was further carried out among those patients without known disease-causing gene mutations. We simultaneously performed a retrospective study on the reported patients exhibiting HSP in other Chinese cohorts. All clinical and molecular characterization from above two groups of Chinese HSP patients were analyzed and summarized. Eventually, we further validated the cellular changes in fibroblasts of two major spastic paraplegia (SPG) patients (SPG4 and SPG11) in vitro. RESULTS: Most patients of ADHSP (94%) are pure forms, whereas most patients of ARHSP (78%) tend to be complicated forms. In ADHSP, we found that SPG4 (79%) was the most prevalent, followed by SPG3A (11%), SPG6 (4%) and SPG33 (2%). Subtle mutations were the common genetic cause for SPG4 patients and most of them located in AAA cassette domain of spastin protein. In ARHSP, the most common subtype was SPG11 (53%), followed by SPG5 (32%), SPG35 (6%) and SPG46 (3%). Moreover, haplotype analysis showed a unique haplotype was shared in 14 families carrying c.334C > T (p.R112(*)) mutation in CYP7B1 gene, suggesting the founder effect. Functionally, we observed significantly different patterns of mitochondrial dynamics and network, decreased mitochondrial membrane potential (deltam), increased reactive oxygen species and reduced ATP content in SPG4 fibroblasts. Moreover, we also found the enlargement of LAMP1-positive organelles and abnormal accumulation of autolysosomes in SPG11 fibroblasts. CONCLUSIONS: Our study present a comprehensive clinical spectrum and genetic landscape for HSP in China. We have also provided additional evidences for mitochondrial and autolysosomal-mediated pathways in the pathogenesis of HSP.
Middle East respiratory syndrome coronavirus (MERS-CoV) has represented a human health threat since 2012. Although several MERS-related CoVs that belong to the same species as MERS-CoV have been identified from bats, they do not use the MERS-CoV receptor, dipeptidyl peptidase 4 (DPP4). Here, we screened 1,059 bat samples from at least 30 bat species collected in different regions in south China and identified 89 strains of lineage C betacoronaviruses, including Tylonycteris pachypus coronavirus HKU4, Pipistrellus pipistrelluscoronavirus HKU5, and MERS-related CoVs. We sequenced the full-length genomes of two positive samples collected from the great evening bat, Ia io, from Guangdong Province. The two genomes were highly similar and exhibited genomic structures identical to those of other lineage C betacoronaviruses. While they exhibited genome-wide nucleotide identities of only 75.3 to 81.2% with other MERS-related CoVs, their gene-coding regions were highly similar to their counterparts, except in the case of the spike proteins. Further protein-protein interaction assays demonstrated that the spike proteins of these MERS-related CoVs bind to the receptor DPP4. Recombination analysis suggested that the newly discovered MERS-related CoVs have acquired their spike genes from a DPP4-recognizing bat coronavirus HKU4. Our study provides further evidence that bats represent the evolutionary origins of MERS-CoV.IMPORTANCE Previous studies suggested that MERS-CoV originated in bats. However, its evolutionary path from bats to humans remains unclear. In this study, we discovered 89 novel lineage C betacoronaviruses in eight bat species. We provide evidence of a MERS-related CoV derived from the great evening bat that uses the same host receptor as human MERS-CoV. This virus also provides evidence for a natural recombination event between the bat MERS-related CoV and another bat coronavirus, HKU4. Our study expands the host ranges of MERS-related CoV and represents an important step toward establishing bats as the natural reservoir of MERS-CoV. These findings may lead to improved epidemiological surveillance of MERS-CoV and the prevention and control of the spread of MERS-CoV to humans.
Title: Palmitic Acid Methyl Ester and Its Relation to Control of Tone of Human Visceral Arteries and Rat Aortas by Perivascular Adipose Tissue Wang N, Kuczmanski A, Dubrovska G, Gollasch M Ref: Front Physiol, 9:583, 2018 : PubMed
Background: Perivascular adipose tissue (PVAT) exerts anti-contractile effects on visceral arteries by release of various perivascular relaxing factors (PVRFs) and opening voltage-gated K(+) (Kv) channels in vascular smooth muscle cells (VSMCs). Palmitic acid methyl ester (PAME) has been proposed as transferable PVRF in rat aorta. Here, we studied PVAT regulation of arterial tone of human mesenteric arteries and clarified the contribution of Kv channels and PAME in the effects. Methods: Wire myography was used to measure vasocontractions of mesenteric artery rings from patients undergoing abdominal surgery. Isolated aortic rings from Sprague-Dawley rats were studied for comparison. PVAT was either left intact or removed from the arterial rings. Vasocontractions were induced by external high K(+) (60 mM), serotonin (5-HT) or phenylephrine. PAME (10 nM-3 muM) was used as vasodilator. Kv channels were blocked by XE991, a Kv7 (KCNQ) channel inhibitor, or by 4-aminopyridine, a non-specific Kv channel inhibitor. PAME was measured in bathing solutions incubated with rat peri-aortic or human visceral adipose tissue. Results: We found that PVAT displayed anti-contractile effects in both human mesenteric arteries and rat aortas. The anti-contractile effects were inhibited by XE991 (30 muM). PAME (EC50 ~1.4 muM) was capable to produce relaxations of PVAT-removed rat aortas. These effects were abolished by XE991 (30 muM), but not 4-aminopyridine (2 mM) or NDGA (10 muM), a lipoxygenases inhibitor. The cytochrome P450 epoxygenase inhibitor 17-octadecynoic acid (ODYA 10 muM) and the soluble epoxide hydrolase inhibitor 12-(3-adamantan-1-ylureido)-dodecanoic acid (AUDA 10 muM) slightly decreased PAME relaxations. PAME up to 10 muM failed to induce relaxations of PVAT-removed human mesenteric arteries. 5-HT induced endogenous PAME release from rat peri-aortic adipose tissue, but not from human visceral adipose tissue. Conclusions: Our data also suggest that Kv7 channels are involved in the anti-contractile effects of PVAT on arterial tone in both rat aorta and human mesenteric arteries. PAME could contribute to PVAT relaxations by activating Kv7 channels in rat aorta, but not in human mesenteric arteries.
A series of 2-arylethenyl-N-methylquinolinium derivatives were designed and synthesized based on our previous research of 2-arylethenylquinoline analogues as multifunctional agents for the treatment of Alzheimer's disease (AD) (Eur. J. Med. Chem. 2015, 89, 349-361). The results of in vitro biological activity evaluation, including beta-amyloid (Abeta) aggregation inhibition, cholinesterase inhibition, and antioxidant activity, showed that introduction of N-methyl in quinoline ring significantly improved the anti-AD potential of compounds. The optimal compound, compound a12, dramatically attenuated the cell death of glutamate-induced HT22 cells by preventing the generation of ROS and increasing the level of GSH. Most importantly, intragastric administration of a12*HAc was well tolerated at doses up to 2000 mg/kg and could traverse blood-brain barrier.
Although the prevalence of Intracytoplasmic sperm injection (ICSI) has increased year by year, there remains concern about the safety of these procedures because of reports of the increased risk for imprinting disorders. Previous research has demonstrated that gonadotropin stimulation contributes to an increased incidence of epimutations in ICSI-derived mice. However, the epimutations in ICSI offspring after removing the effect of gonadotropin stimulation and the possibility that epimutations are reversible by developmental reprogramming has not been investigated. Our study is the first to investigate the effect of ICSI itself on methylation and exclude the effect of superovulation using the kidney tissues from the adult and old mice. We found reduced methylation and up-regulated expression of the imprinted genes, H19, Mest and Peg3, in adult ICSI mice, but the above alterations observed in adult mice were not detected in old ICSI mice. At the Snrpn DMR, methylation status was not altered in adult ICSI-derived mice, but hypermethylation and correlated down-regulated expression of Snrpn were observed in old mice. In conclusion, ICSI manipulation and early embryo culture resulted in alterations of methylation in differentially methylated region of H19, Mest, Peg3 and Snrpn, and the alterations were reprogrammed by developmental reprogramming.
Title: Immobilization of Aspergillus terreus lipase in self-assembled hollow nanospheres for enantioselective hydrolysis of ketoprofen vinyl ester Hu C, Wang N, Zhang W, Zhang S, Meng Y, Yu X Ref: J Biotechnol, 194:12, 2015 : PubMed
The aim of this study was to improve the ability of Aspergillus terreus lipase to separate the racemic ketoprofen vinyl ester into individual enantiomers using hollow self-assembly alginate-graft-poly(ethylene glycol)/alpha-cyclodextrins (Alg-g-PEG/alpha-CD) spheres as enzyme immobilization carriers. The morphology and size of the Alg-g-PEG/alpha-CD particles were investigated by transmission electron microscopy (TEM) and were found to be nanoscale. To facilitate recycling, calcium alginate (CA) beads were developed to encapsulate Alg-g-PEG/alpha-CD particles, thereby producing Alg-g-PEG/alpha-CD/CA composite beads. The influence of buffer pH and enzyme concentration during immobilization was studied along with the biocatalyst's kinetic parameters. When the immobilized enzyme was under optimal conditions in the resolution reaction, maximal conversion (approximately 45.9%) and enantioselectivity (approximately 128.8) were obtained. The immobilized A. terreus lipase maintained excellent performance even after 20 reuses and retained nearly 100% of its original activity after 24 weeks of storage at 4 degrees C.
Genetic sex determination by W and Z chromosomes has developed independently in different groups of organisms. To better understand the evolution of sex chromosomes and the plasticity of sex-determination mechanisms, we sequenced the whole genomes of a male (ZZ) and a female (ZW) half-smooth tongue sole (Cynoglossus semilaevis). In addition to insights into adaptation to a benthic lifestyle, we find that the sex chromosomes of these fish are derived from the same ancestral vertebrate protochromosome as the avian W and Z chromosomes. Notably, the same gene on the Z chromosome, dmrt1, which is the male-determining gene in birds, showed convergent evolution of features that are compatible with a similar function in tongue sole. Comparison of the relatively young tongue sole sex chromosomes with those of mammals and birds identified events that occurred during the early phase of sex-chromosome evolution. Pertinent to the current debate about heterogametic sex-chromosome decay, we find that massive gene loss occurred in the wake of sex-chromosome 'birth'.
Hirsutella minnesotensis [Ophiocordycipitaceae (Hypocreales, Ascomycota)] is a dominant endoparasitic fungus by using conidia that adhere to and penetrate the secondary stage juveniles of soybean cyst nematode. Its genome was de novo sequenced and compared with five entomopathogenic fungi in the Hypocreales and three nematode-trapping fungi in the Orbiliales (Ascomycota). The genome of H. minnesotensis is 51.4 Mb and encodes 12,702 genes enriched with transposable elements up to 32%. Phylogenomic analysis revealed that H. minnesotensis was diverged from entomopathogenic fungi in Hypocreales. Genome of H. minnesotensis is similar to those of entomopathogenic fungi to have fewer genes encoding lectins for adhesion and glycoside hydrolases for cellulose degradation, but is different from those of nematode-trapping fungi to possess more genes for protein degradation, signal transduction, and secondary metabolism. Those results indicate that H. minnesotensis has evolved different mechanism for nematode endoparasitism compared with nematode-trapping fungi. Transcriptomics analyses for the time-scale parasitism revealed the upregulations of lectins, secreted proteases and the genes for biosynthesis of secondary metabolites that could be putatively involved in host surface adhesion, cuticle degradation, and host manipulation. Genome and transcriptome analyses provided comprehensive understanding of the evolution and lifestyle of nematode endoparasitism.
Tanshinol (3-(3',4'-dihydroxyphenyl)-(2R)-lactic acid, TSL) is widely used in traditional Chinese medicine for the treatment of cardiovascular and cerebrovascular diseases. Here, we assessed whether TSL protected hippocampus and attenuated vascular dementia (VD) development in rats. The behavioral analysis showed that TSL could decrease the distance and latency time, and increase the swim speed in water maze in rats subjected to VD. TSL remarkably increased acetylcholine level and decreased acetylcholinesterase activity in rats subjected to VD. Likewise, TSL remarkably decreased malondialdehyde and increased superoxide dismutase levels in rats subjected to VD. Furthermore, treatment with TSL reduced the level of dead neurons in dentate gyrus. In addition, TSL upregulated growth-associated protein 43 (GAP43) and vascular endothelial growth factor (VEGF) expression and downregulated phosphorylated Akt (p-AKt) and phosphorylated glycogen synthase kinase (p-GSK3beta) expression in hippocampus in rats subjected to VD. These results suggest that TSL may be a potential compound in VD model.
Title: Identification of residues on human receptor DPP4 critical for MERS-CoV binding and entry Song W, Wang Y, Wang N, Wang D, Guo J, Fu L, Shi X Ref: Virology, 471-473C:49, 2014 : PubMed
Middle East respiratory syndrome coronavirus (MERS-CoV) infects host cells through binding the receptor binding domain (RBD) on its spike glycoprotein to human receptor dipeptidyl peptidase 4 (hDPP4). Here, we report identification of critical residues on hDPP4 for RBD binding and virus entry through analysis of a panel of hDPP4 mutants. Based on the RBD-hDPP4 crystal structure we reported, the mutated residues were located at the interface between RBD and hDPP4, which potentially changed the polarity, hydrophobic or hydrophilic properties of hDPP4, thereby interfering or disrupting their interaction with RBD. Using surface plasmon resonance (SPR) binding analysis and pseudovirus infection assay, we showed that several residues in hDPP4-RBD binding interface were important on hDPP4-RBD binding and viral entry. These results provide atomic insights into the features of interactions between hDPP4 and MERS-CoV RBD, and also provide potential explanation for cellular and species tropism of MERS-CoV infection.
Title: Effects of thyroxin and donepezil on hippocampal acetylcholine content and syntaxin-1 and munc-18 expression in adult rats with hypothyroidism Wang N, Cai Y, Wang F, Zeng X, Jia X, Tao F, Zhu D Ref: Exp Ther Med, 7:529, 2014 : PubMed
Adult-onset hypothyroidism induces various impairments in hippocampus-dependent cognitive function, in which numerous synaptic proteins and neurotransmitters are involved. Donepezil (DON), an acetylcholinesterase inhibitor, has been shown to be efficient in improving cognitive function. The aim of the present study was to investigate the effects of adult-onset hypothyroidism on the expression levels of the synaptic proteins syntaxin-1 and munc-18, as well as the content of the neurotransmitter acetylcholine (ACh) in the hippocampus. In addition, the study explored the effects of thyroxin (T4) and DON treatment on the altered parameters. The study involved 55 Sprague-Dawley rats that were randomly divided into five groups: Control, hypothyroid (0.05% 6-n-propyl-2-thiouracil; added to the drinking water), hypothyroid treated with T4 (6 mug/100 g body weight once daily; intraperitoneal injection), hypothyroid treated with DON (0.005%; added to the drinking water) and hypothyroid treated with a combination of the two drugs (6 mug/100 g T4 and 0.005% DON). The concentration of ACh was determined in the homogenized hippocampus of each animal by alkaline hydroxylamine colorimetry. The protein levels of syntaxin-1 and munc-18 were determined by immunohistochemistry. The results showed that the content of ACh in the hippocampi of the hypothyroid rats was significantly decreased compared with that in the controls and that T4 monotherapy and DON administration restored the ACh content to normal values. In the hippocampi of the hypothyroid group, munc-18 was expressed at significantly lower levels, while the expression levels of syntaxin-1 were increased compared with the levels in the control group. Treatment with T4 alone restored the expression of syntaxin-1 but failed to normalize munc-18 expression levels. The co-administration of T4 and DON returned the munc-18 levels to normal values. These observations indicate that adult-onset hypothyroidism induces alterations in the levels of munc-18, syntaxin-1 and ACh in the hippocampus. Syntaxin-1 and ACh levels were restored by T4 monotherapy while munc-18 levels were not. In addition, the co-administration of T4 and DON resulted in more effective restoration than either alone. The thyroid hormone has a direct effect on metabolism of hippocampal ACh in adult rats and DON is helpful for treatment of synaptic protein impairment induced by hypothyroidism.
Title: Draft Genome Sequence of the Bioelectricity-Generating and Dye-Decolorizing Bacterium Proteus hauseri Strain ZMd44 Wang N, Ng IS, Chen PT, Li Y, Chen YC, Chen BY, Lu Y Ref: Genome Announc, 2:e00992, 2014 : PubMed
Proteus hauseri ZMd44 (CGMCC 6746), as a crucial biodecolorizing, bioelectricity-generating, and copper-resistant bacterium, is distinguished from the urinary pathogens Proteus penneri and Proteus mirabilis. To further investigate the genetic functions of this strain, the genome sequence and annotation of its open reading frames, which consist of 3,875,927 bp (G+C content, 38.12%), are presented here.
Title: Effects of additives on lipase immobilization in microemulsion-based organogels Zhang WW, Wang N, Zhang L, Wu WX, Hu CL, Yu XQ Ref: Appl Biochem Biotechnol, 172:3128, 2014 : PubMed
An inexpensive, facile, and environmentally benign method was developed to improve the activity and stability of Candida rugosa lipase (triacylglycerol acylhydrolase) immobilized on microemulsion-based organogels (CRL MBGs) via the addition of additives during immobilization. The additives used were polyethylene glycol (PEG) or polysaccharides. This study is the first report on the effect of additives in CRL MBGs. Among the tested additives, PEG produced the most improvement in the immobilized CRL, enhancing its stability in organic solvents (specifically polar solvents). The results of circular dichroism and fluorescence spectra experiments indicated that exposure of the acidic CRL to electronegative additives in the buffer, such as polyethylenimine and the electropositive surfactant cetyltrimethylammonium bromide, may change the lipase secondary structure, ultimately causing enzyme inactivation. However, sodium bis(2-ethylhexyl)sulfosuccinate and PEG 2000 had minimal effects on the secondary structure of CRL. The CRL MBGs containing PEG 2000 demonstrated remarkable retention of their catalytic activity during the recycling test. No significant changes in enzymatic activity were observed, even after nine runs, and 90 % of the original yield was maintained after 15 cycles.
Title: An ultra-sensitive acetylcholinesterase biosensor based on reduced graphene oxide-Au nanoparticles-beta-cyclodextrin/Prussian blue-chitosan nanocomposites for organophosphorus pesticides detection Zhao H, Ji X, Wang B, Wang N, Li X, Ni R, Ren J Ref: Biosensors & Bioelectronics, 65C:23, 2014 : PubMed
This work reports a novel, ultrasensitive, and selective sensing platform based on a direct electrodeposition of electrochemical reduced graphene oxide (ERGO)-Au nanoparticles (AuNPs)-beta-cyclodextrin (beta-CD) and Prussian blue-chitosan (PB-CS) on glass carbon electrode (GCE) for efficiently fixed acetylcholinesterase (AChE) to fabricate organophosphorus pesticides (OPs) biosensor. The PB-CS not only effectively catalyzed the oxidation of thiocholine (TCh), but also shifted its oxidation potential from 0.68 to 0.2V, and accordingly the sensitivity of the biosensor was obviously improved. The synergistic effect between ERGO and AuNPs significantly promoted the electron transfer between PB and GCE, and remarkably enhanced the electrochemical oxidation of TCh. Besides, beta-CD could interact with substrate by reversible bonding, which is contribute to increase the enrichment of the substrate and improve the selectivity and sensitivity of the biosensor. The integration of ERGO-AuNPs-beta-CD with PB-CS provided an advantageous and high-performance platform for sensing applications. Based on the inhibition of OPs on AChE activity, the sensor showed wide linear ranges of 7.98-2.00x103pgmL-1 and 4.3-1.00x103pgmL-1 with low detection limits of 4.14pgmL-1 and 1.15pgmL-1 for malathion and carbaryl, respectively. The proposed biosensor exhibited short response time, good stability and high sensitivity, which can be used for direct analysis of practical samples.
Title: Complete Genome Sequence of Xanthomonas citri subsp. citri Strain Aw12879, a Restricted-Host-Range Citrus Canker-Causing Bacterium Jalan N, Kumar D, Yu F, Jones JB, Graham JH, Wang N Ref: Genome Announc, 1:, 2013 : PubMed
Xanthomonas citri subsp. citri causes citrus canker. The Asiatic strain has a broad host range, whereas the Wellington variant has a restricted host range. Here, we present the complete genome of X. citri subsp. citri strain A(W)12879. This study lays the foundation to further characterize the mechanisms for virulence and host range of X. citri.
The spike glycoprotein (S) of recently identified Middle East respiratory syndrome coronavirus (MERS-CoV) targets the cellular receptor, dipeptidyl peptidase 4 (DPP4). Sequence comparison and modeling analysis have revealed a putative receptor-binding domain (RBD) on the viral spike, which mediates this interaction. We report the 3.0 A-resolution crystal structure of MERS-CoV RBD bound to the extracellular domain of human DPP4. Our results show that MERS-CoV RBD consists of a core and a receptor-binding subdomain. The receptor-binding subdomain interacts with DPP4 beta-propeller but not its intrinsic hydrolase domain. MERS-CoV RBD and related SARS-CoV RBD share a high degree of structural similarity in their core subdomains, but are notably divergent in the receptor-binding subdomain. Mutagenesis studies have identified several key residues in the receptor-binding subdomain that are critical for viral binding to DPP4 and entry into the target cell. The atomic details at the interface between MERS-CoV RBD and DPP4 provide structural understanding of the virus and receptor interaction, which can guide development of therapeutics and vaccines against MERS-CoV infection.
Title: Prenatal nicotine exposure alters lung function and airway geometry through alpha7 nicotinic receptors Wongtrakool C, Wang N, Hyde DM, Roman J, Spindel ER Ref: American Journal of Respiratory Cellular & Molecular Biology, 46:695, 2012 : PubMed
Maternal smoking during pregnancy has been associated with adverse effects on respiratory health. Whereas the epidemiologic link is incontrovertible, the mechanisms responsible for this association are still poorly understood. Although cigarette smoke has many toxic constituents, nicotine, the major addictive component in cigarette smoke, may play a more significant role than previously realized. The objectives of this study were to determine whether exposure to nicotine prenatally leads to alterations in pulmonary function and airway geometry in offspring, and whether alpha7 nicotinic acetylcholine receptors (nAChRs) mediate these effects. In a murine model of in utero nicotine exposure, pulmonary function, airway size and number, methacholine response, and collagen deposition were examined. Exposure periods included Gestation Days 7-21, Gestation Day 14 to Postnatal Day 7, and Postnatal Days 3-15. Prenatal nicotine exposure decreases forced expiratory flows in offspring through alpha7 nAChR-mediated signals, and the critical period of nicotine exposure was between Prenatal Day 14 and Postnatal Day 7. These physiologic changes were associated with increased airway length and decreased diameter. In addition, adult mice exposed to prenatal nicotine exhibit an increased response to methacholine challenge, even in the absence of allergic sensitization. Collagen expression was increased between adjacent airways and vessels, which was absent in alpha7 nAChR knockout mice. These observations provide a unified mechanism of how maternal smoking during pregnancy may lead to lifelong alterations in offspring pulmonary function and increased risk of asthma, and suggest potential targets to counteract those effects.
We report a draft sequence of the genome of Gordonia neofelifaecis NRRL B-59395, a cholesterol-degrading actinomycete isolated from fresh feces of a clouded leopard (Neofelis nebulosa). As predicted, the reported genome contains several gene clusters for cholesterol degradation. This is the second available genome sequence of the family Gordoniaceae.
Title: Comparative genomic analysis of Xanthomonas axonopodis pv. citrumelo F1, which causes citrus bacterial spot disease, and related strains provides insights into virulence and host specificity Jalan N, Aritua V, Kumar D, Yu F, Jones JB, Graham JH, Setubal JC, Wang N Ref: Journal of Bacteriology, 193:6342, 2011 : PubMed
Xanthomonas axonopodis pv. citrumelo is a citrus pathogen causing citrus bacterial spot disease that is geographically restricted within the state of Florida. Illumina, 454 sequencing, and optical mapping were used to obtain a complete genome sequence of X. axonopodis pv. citrumelo strain F1, 4.9 Mb in size. The strain lacks plasmids, in contrast to other citrus Xanthomonas pathogens. Phylogenetic analysis revealed that this pathogen is very close to the tomato bacterial spot pathogen X. campestris pv. vesicatoria 85-10, with a completely different host range. We also compared X. axonopodis pv. citrumelo to the genome of citrus canker pathogen X. axonopodis pv. citri 306. Comparative genomic analysis showed differences in several gene clusters, like those for type III effectors, the type IV secretion system, lipopolysaccharide synthesis, and others. In addition to pthA, effectors such as xopE3, xopAI, and hrpW were absent from X. axonopodis pv. citrumelo while present in X. axonopodis pv. citri. These effectors might be responsible for survival and the low virulence of this pathogen on citrus compared to that of X. axonopodis pv. citri. We also identified unique effectors in X. axonopodis pv. citrumelo that may be related to the different host range as compared to that of X. axonopodis pv. citri. X. axonopodis pv. citrumelo also lacks various genes, such as syrE1, syrE2, and RTX toxin family genes, which were present in X. axonopodis pv. citri. These may be associated with the distinct virulences of X. axonopodis pv. citrumelo and X. axonopodis pv. citri. Comparison of the complete genome sequence of X. axonopodis pv. citrumelo to those of X. axonopodis pv. citri and X. campestris pv. vesicatoria provides valuable insights into the mechanism of bacterial virulence and host specificity.
As an obligatory parasite of humans, the body louse (Pediculus humanus humanus) is an important vector for human diseases, including epidemic typhus, relapsing fever, and trench fever. Here, we present genome sequences of the body louse and its primary bacterial endosymbiont Candidatus Riesia pediculicola. The body louse has the smallest known insect genome, spanning 108 Mb. Despite its status as an obligate parasite, it retains a remarkably complete basal insect repertoire of 10,773 protein-coding genes and 57 microRNAs. Representing hemimetabolous insects, the genome of the body louse thus provides a reference for studies of holometabolous insects. Compared with other insect genomes, the body louse genome contains significantly fewer genes associated with environmental sensing and response, including odorant and gustatory receptors and detoxifying enzymes. The unique architecture of the 18 minicircular mitochondrial chromosomes of the body louse may be linked to the loss of the gene encoding the mitochondrial single-stranded DNA binding protein. The genome of the obligatory louse endosymbiont Candidatus Riesia pediculicola encodes less than 600 genes on a short, linear chromosome and a circular plasmid. The plasmid harbors a unique arrangement of genes required for the synthesis of pantothenate, an essential vitamin deficient in the louse diet. The human body louse, its primary endosymbiont, and the bacterial pathogens that it vectors all possess genomes reduced in size compared with their free-living close relatives. Thus, the body louse genome project offers unique information and tools to use in advancing understanding of coevolution among vectors, symbionts, and pathogens.
This study examined the advantages of the use of biomarkers as an early warning system by applying it to different shrimp farming systems in Soctrang and Camau provinces, main shrimp producers in Mekong River Delta, Vietnam. Shrimp were collected at 15 different farms divided into four different farming systems: three farms were converted from originally rice paddies into intensive shrimp farming systems (IS1, IS2, IS3); three farms were rice-shrimp integrated farming systems (RS4, RS5, RS6); three farms were intensive farming systems (IS7, IS8, IS9); six farms were extensive shrimp farming systems (From ES1 to ES6). Lipid peroxidation (LPO) and total glutathione (GSH) were measured as well as catalase (CAT), glutathione peroxidase (GPX), glutathione S-transferase (GST) and acetylcholinesterase activities (ACHE). Organ specificity was observed between gills and hepatopancreas with generally higher activity of GST in gills (GSTG) whereas the contrary was observed for LPO level in gills (LPOG). Hierarchical clustering and principal component analysis clearly indicated that shrimp reared in extensive culture system formed a distinct group from those reared in intensive or rice-shrimp integrated systems. CAT in gills (CATG), GPX in gills (GPXG) and hepatopancreas (GPXHP) and ACHE in muscle (ACHEM) of shrimp collected in extensive farms showed a general higher level than those in intensively farmed shrimp. On the contrary, we observed clear high levels of GSTG and GST in hepatopancreas (GSTHP) and LPOG and hepatopancreas (LPOHP) of shrimp sampled in intensive and rice-shrimp integrated systems. Thus, we propose that LPO and CAT, GPX, GST and ACHE can be used as a set of biomarkers for the assessment of health condition and can discriminate between shrimp cultivated in different farming systems. These findings provide the usefulness of integrating a set of biomarkers to define the health status of shrimp in different shrimp culture systems.
Title: Biocatalytic synthesis and in vitro release of biodegradable linear polyesters with pendant ketoprofen Wang HY, Zhang WW, Wang N, Li C, Li K, Yu XQ Ref: Biomacromolecules, 11:3290, 2010 : PubMed
Enzyme-catalyzed polycondensation for the synthesis of polyester prodrugs of ketoprofen was reported. Lipase acrylic resin from Candida antarctica (CAL-B) was used to synthesize the linear polyesters with pendent ketoprofen groups based on ketoprofen glycerol ester, poly(ethylene glycol), and divinyl sebacate. The products were characterized by GPC and (1)H NMR. The results indicated that the molecular weight and yields of the polyesters depend on experimental conditions such as temperature and feed ratio. The in vitro study showed that the drug release from the polyester was slow under physiological conditions, which indicated that the polyester could be a promising prodrug with extended pharmacological effects by delayed release of ketoprofen.
Rg1 and Rb1 are two major active compounds of ginseng that facilitate learning and memory. The present study aimed to compare the nootropic effects of Rg1 and Rb1 in a scopolamine induced dementia mice model. After 6 and 12 mg/kg of Rg1 and Rb1 intraperitoneal administration to mice for 7 days, their effects were assessed using the step-down passive avoidance (SD) and the Morris water maze (MWM) tests, the acetylcholinesterase (AChE) activity, acetylcholine (ACh) content and serotonin (5-HT) level in the hippocampus were analysed after SD and MWM tests. The results showed that Rg1 and Rb1 ameliorated cognition-deficiency in mice with dementia. Rg1 showed stronger effects than Rb1 on escape acquisition in MWM. Both Rg1 and Rb1 increased ACh levels in the hippocampus, but Rg1 inhibited AChE activity while Rb1 had no effect on AChE activity. Both Rg1 and Rb1 inhibited the decrease of 5-HT induced by scopolamine, but Rb1 was more active than the same dose of Rg1. These results demonstrate that multiple administrations of Rg1 and Rb1 are effective in improving memory deficiency induced by scopolamine. Rg1 appears to be more potent than Rb1 in improving acquisition impairment, and the two ginsenosides may act through different mechanisms.
Title: The positive allosteric modulator morantel binds at noncanonical subunit interfaces of neuronal nicotinic acetylcholine receptors Seo S, Henry JT, Lewis AH, Wang N, Levandoski MM Ref: Journal of Neuroscience, 29:8734, 2009 : PubMed
We are interested in the positive allosteric modulation of neuronal nicotinic acetylcholine (ACh) receptors and have recently shown that the anthelmintic compound morantel potentiates by enhancing channel gating of the alpha3beta2 subtype. Based on the demonstration that morantel-elicited currents were inhibited by the classic ACh competitor dihydro-beta-erythroidine in a noncompetitive manner and that morantel still potentiates at saturating concentrations of agonist (Wu et al., 2008), we hypothesized that morantel binds at the noncanonical beta2(+)/alpha3(-) subunit interface. In the present study, we created seven cysteine-substituted subunits by site-directed mutagenesis, choosing residues in the putative morantel binding site with the aid of structural homology models. We coexpressed the mutant subunits and their respective wild-type partners in Xenopus oocytes and characterized the morantel potentiation of ACh-evoked currents, as well as morantel-evoked currents, before and after treatment with a variety of methanethiosulfonate (MTS)-based compounds, using voltage-clamp recordings. The properties of four of the seven mutants, two residues on each side of the interface, were changed by MTS treatments. Coapplication with ACh enhanced the extent of MTS modification for alpha3A106Cbeta2 and alpha3beta2S192C receptors. The activities of two mutants, alpha3T115Cbeta2 and alpha3beta2T150C, were dramatically altered by MTS modification. For alpha3beta2T150C, while peak current amplitudes were reduced, potentiation was enhanced. For alpha3T115Cbeta2, both current amplitudes and potentiation were reduced. MTS modification and morantel were mutually inhibitory: MTS treatment decreased morantel-evoked currents and morantel decreased the rate of MTS modification. We conclude that the four residues showing MTS effects contribute to the morantel binding site.
Title: Markedly enhancing lipase-catalyzed synthesis of nucleoside drugs' ester by using a mixture system containing organic solvents and ionic liquid Liu BK, Wang N, Chen ZC, Wu Q, Lin XF Ref: Bioorganic & Medicinal Chemistry Lett, 16:3769, 2006 : PubMed
Eightfold higher yields and three times faster reaction rates were achieved by means of using a mixture solvent system composed of 90% acetone and 10% [BMIM]BF4 in the lipase-catalyzed regioselective synthesis of polymerizable ester of nucleoside drugs.
Title: Hidden function of neuronal nicotinic acetylcholine receptor beta2 subunits in ganglionic transmission: comparison to alpha5 and beta4 subunits Wang N, Orr-Urtreger A, Chapman J, Ergun Y, Rabinowitz R, Korczyn AD Ref: Journal of Neurology Sci, 228:167, 2005 : PubMed
Neuronal nicotinic acetylcholine receptors (nAChR), which modulate fast excitatory postsynaptic potentials (f-EPSP), are located on both pre- and postganglionic sites in the autonomic nervous system (ANS). The receptor subunits alpha3, alpha5, alpha7, beta2 and beta4 are present in autonomic ganglia in various combinations and modulate acetylcholine (ACh) transmission. In the present study, autonomic functions were systemically examined in mice lacking beta2 subunits (beta2-/-) to further understand the functional role of beta2 subunits in modulating ganglionic transmission. The results show normal autonomic functions, both under physiological conditions and in perturbed conditions, on thermoregulation, pupillary size, heart rate responses and ileal contractile reactions. This suggests that the function of beta2-containing receptors in ganglionic transmission is hidden by the predominant beta4 containing receptors and confirms previous studies which suggest that alpha3alpha5beta4 nAChRs are sufficient for autonomic transmission. On the other hand, beta2-containing receptors have only a minor function on postsynaptic responses to ACh, but may modulate ACh release presynaptically, although there is no evidence for this.
Neuronal nicotinic acetylcholine receptors (nAChRs) are composed of 12 subunits (alpha2-alpha10 and beta2-beta4). alpha5 Subunits, expressed throughout the central nervous system (CNS) and the autonomic nervous system (ANS), possess unique pharmacological properties. The effects of oxotremorine (OXO) on autonomic functions and tremor were examined in mice lacking alpha5 nAChR subunits (alpha5-/-) and compared with those in wild-type (WT) control mice. The alpha5-/- mice showed significantly increased salivation and tremor responses to OXO. The hypothermia, bradycardia and defecation induced by OXO were of similar magnitudes in the two mouse strains. The enhanced OXO effects in alpha5-/- mice indicate inhibitory effects of alpha5 subunits in autonomic ganglia, and support the participation of these subunits in cholinergic transmission in autonomic ganglia.
Title: Autonomic function of neuronal nicotinic acetylcholine receptor subunits alpha5, beta2 and beta4. Wang N, Orr-Urtreger A, Korczyn AD Ref: Cholinergic Mechanisms, CRC Press, :317, 2004 : PubMed
Neuronal nicotinic acetylcholine receptors (nAChR) are composed of 12 subunits (alpha 2-alpha 10 and beta 2-beta 4), which play the central role in autonomic transmission. beta 4 subunits are abundantly expressed in autonomic ganglia, forming acetylcholine binding sites and ion channels with alpha 3 or alpha 3 and alpha 5 subunits as pentameric receptors. To investigate the physiological and pharmacological properties of beta 4 subunits in autonomic ganglia, we measured autonomic functions in knockout mice lacking nAChR subunit beta 4 (beta 4(-/-)) and wild-type mice. beta 4(-/-) mice had an attenuated bradycardiac response to high frequency (60 pulse/s) vagal stimulation, as well as an increased sensitivity to hexamethonium blockade at low dose (3 mg/kg) and a reduced ileal contractile response to the nicotinic agonists cytisine, dimethylphenylpiperazinium iodide, nicotine (10 mg/kg each), and epibatidine (0.1 mg/kg). The results suggest that beta 4 subunits are important components of nAChRs in autonomic ganglia. Deficiency of beta 4 subunits altered ion channel properties, conductance, and sensitivity and affinity of receptors to agonists and antagonists, affecting ganglionic transmission.
AIM To investigate the possible association of microsomal epoxide hydrolase (mEH) Tyr113His polymorphism with susceptibility to esophageal squamous cell carcinoma (ESCC) in a population of North China.
METHODS:
The mEH Tyr113His genotypes were determined by polymerase-chain reaction (PCR)-restriction fragment length polymorphism (RFLP) analysis in 257 patients with esophageal squamous cell carcinoma (ESCC) and 252 healthy subjects as a control group.
RESULTS:
The frequencies for Tyr and His alleles were 44.2%, 55.8% in ESCC patients, and 44.0% and 56.0% in healthy subjects, respectively. No statistic difference in allele distribution was observed between ESCC patients and controls (chi2=0.008, P=0.929). The overall genotype distribution difference was not observed between cancer cases and controls (chi2=2.116, P=0.347). Compared with Tyr/Tyr genotype, neither His/His genotype nor in combination with Tyr/His genotype significantly modified the risk of the development of ESCC, the adjusted odds ratio was 1.076 (95% CI=0.850-1.361) and 0.756 (95% CI=0.493-1.157), respectively. When stratified for sex, age, smoking status and family history of upper gastrointestinal cancer, His/His genotype alone or in combination with Tyr/His genotype also did not show any significant influence on the risk of developing ESCC.
CONCLUSION:
MEH Tyr113His polymorphism may not be used as a stratification marker in screening individuals at a high risk of ESCC.
Neuronal acetylcholine nicotinic receptors (nAChR) are composed of 12 subunits (alpha2-10, beta2-4), of which alpha3, alpha5, alpha7, beta2 and beta4 subunits are known to exist in the autonomic nervous system (ANS). alpha5 subunits possess unique biophysical and pharmacological properties. The present study was undertaken to examine the functional role and pharmacological properties of the nAChR alpha5 subunits in the ANS using mice lacking alpha5 nAChR subunits (alpha5-/-). These mice grew to normal size showing no obvious physical or neurological deficit. They also showed normality in thermoregulation, pupil size and resting heart rate under physiological conditions. The heart rate and rectal temperature did not differ between alpha5-/- and wild-type mice during exposure to cold stress. An impairment of cardiac parasympathetic ganglionic transmission was observed during high frequency vagal stimulation, which caused cardiac arrest in all wild-type animals while alpha5-/- mice were more resistant. Deficiency of alpha5 subunits strikingly increased the sensitivity to a low concentration of hexamethonium, leading to a nearly complete blockade of bradycardia in response to vagal stimulation. Such a concentration of hexamethonium only slightly depressed the effects of vagal stimulation in control mice. Deficiency of alpha5 subunits significantly increased ileal contractile responses to cytisine and epibatidine. These results suggest that alpha5 subunits may affect the affinity and sensitivity of agonists and antagonists in the native receptors. Previous studies revealed that alpha5 subunits form functional receptors only in combination with other alpha and beta subunits. Thus, the data presented here imply that alpha5 subunits modulate the activity of nAChR in autonomic ganglia in vivo.
Title: The role of neuronal nicotinic acetylcholine receptor subunits in autonomic ganglia: lessons from knockout mice Wang N, Orr-Urtreger A, Korczyn AD Ref: Prog Neurobiol, 68:341, 2002 : PubMed
Neuronal nicotinic acetylcholine receptors (nAChR), composed of 12 subunits (alpha2-alpha10, beta2-beta4), are expressed in autonomic ganglia, playing a central role in autonomic transmission. The repertoire of nicotinic subunits in autonomic ganglia includes alpha3, alpha5, alpha7, beta2 and beta4 subunits. In the last 10 years, heterologous expression studies have revealed much about the nature of neuronal nAChRs. However, there is only limited understanding of subunit actions in autonomic system. Functional deletions of subunit by gene knockout in animals could overcome these limitations. We review recent studies on nAChRs on autonomic ganglia for physiological and pharmacological properties and potential locations of the subunits.
Title: Poster: Expression of muscarinic receptor subtypes (M1-M5) and adenylate cyclase subtypes II and IV in the human aqueous humor outflow pathway Zhang X, Schroeder A, Wang N, Erickson KA Ref: Life Sciences, 64:591, 1999 : PubMed
