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1.
Mol Cell ; 84(3): 447-462.e10, 2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-38244544

RESUMO

Tumor suppressor BRCA2 functions in homology-directed repair (HDR), the protection of stalled replication forks, and the suppression of replicative gaps, but their relative contributions to genome integrity and chemotherapy response are under scrutiny. Here, we report that mouse and human cells require a RAD51 filament stabilization motif in BRCA2 for fork protection and gap suppression but not HDR. In mice, the loss of fork protection/gap suppression does not compromise genome stability or shorten tumor latency. By contrast, HDR deficiency increases spontaneous and replication stress-induced chromosome aberrations and tumor predisposition. Unlike with HDR, fork protection/gap suppression defects are also observed in Brca2 heterozygous cells, likely due to reduced RAD51 stabilization at stalled forks/gaps. Gaps arise from PRIMPOL activity, which is associated with 5-hydroxymethyl-2'-deoxyuridine sensitivity due to the formation of SMUG1-generated abasic sites and is exacerbated by poly(ADP-ribose) polymerase (PARP) inhibition. However, HDR proficiency has the major role in mitigating sensitivity to chemotherapeutics, including PARP inhibitors.


Assuntos
Proteína BRCA2 , Replicação do DNA , Rad51 Recombinase , Animais , Humanos , Camundongos , Proteína BRCA2/metabolismo , Reparo do DNA , Instabilidade Genômica , Genômica , Rad51 Recombinase/genética , Rad51 Recombinase/metabolismo , Reparo de DNA por Recombinação
2.
Proc Natl Acad Sci U S A ; 121(35): e2320804121, 2024 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-39172790

RESUMO

Breast Cancer Type 1 Susceptibility Protein (BRCA1) is a tumor-suppressor protein that regulates various cellular pathways, including those that are essential for preserving genome stability. One essential mechanism involves a BRCA1-A complex that is recruited to double-strand breaks (DSBs) by RAP80 before initiating DNA damage repair (DDR). How RAP80 itself is recruited to DNA damage sites, however, is unclear. Here, we demonstrate an intrinsic correlation between a methyltransferase DOT1L-mediated RAP80 methylation and BRCA1-A complex chromatin recruitment that occurs during cancer cell radiotherapy resistance. Mechanistically, DOT1L is quickly recruited onto chromatin and methylates RAP80 at multiple lysines in response to DNA damage. Methylated RAP80 is then indispensable for binding to ubiquitinated H2A and subsequently triggering BRCA1-A complex recruitment onto DSBs. Importantly, DOT1L-catalyzed RAP80 methylation and recruitment of BRCA1 have clinical relevance, as inhibition of DOT1L or RAP80 methylation seems to enhance the radiosensitivity of cancer cells both in vivo and in vitro. These data reveal a crucial role for DOT1L in DDR through initiating recruitment of RAP80 and BRCA1 onto chromatin and underscore a therapeutic strategy based on targeting DOT1L to overcome tumor radiotherapy resistance.


Assuntos
Proteína BRCA1 , Reparo do DNA , Chaperonas de Histonas , Histona-Lisina N-Metiltransferase , Humanos , Proteína BRCA1/metabolismo , Proteína BRCA1/genética , Chaperonas de Histonas/metabolismo , Chaperonas de Histonas/genética , Histona-Lisina N-Metiltransferase/metabolismo , Histona-Lisina N-Metiltransferase/genética , Metilação , Linhagem Celular Tumoral , Quebras de DNA de Cadeia Dupla , Cromatina/metabolismo , Metiltransferases/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Animais , Feminino , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Camundongos , Tolerância a Radiação/genética , Metilação de DNA
3.
Mol Cell ; 71(4): 621-628.e4, 2018 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-30057198

RESUMO

FANCA is a component of the Fanconi anemia (FA) core complex that activates DNA interstrand crosslink repair by monoubiquitination of FANCD2. Here, we report that purified FANCA protein catalyzes bidirectional single-strand annealing (SA) and strand exchange (SE) at a level comparable to RAD52, while a disease-causing FANCA mutant, F1263Δ, is defective in both activities. FANCG, which directly interacts with FANCA, dramatically stimulates its SA and SE activities. Alternatively, FANCB, which does not directly interact with FANCA, does not stimulate this activity. Importantly, five other patient-derived FANCA mutants also exhibit deficient SA and SE, suggesting that the biochemical activities of FANCA are relevant to the etiology of FA. A cell-based DNA double-strand break (DSB) repair assay demonstrates that FANCA plays a direct role in the single-strand annealing sub-pathway (SSA) of DSB repair by catalyzing SA, and this role is independent of the canonical FA pathway and RAD52.


Assuntos
Reparo do DNA por Junção de Extremidades , Reparo de Erro de Pareamento de DNA , DNA/genética , Proteína do Grupo de Complementação A da Anemia de Fanconi/genética , Proteína do Grupo de Complementação G da Anemia de Fanconi/genética , Proteínas de Grupos de Complementação da Anemia de Fanconi/genética , Reparo de DNA por Recombinação , Animais , Baculoviridae/genética , Baculoviridae/metabolismo , Linhagem Celular Tumoral , Clonagem Molecular , DNA/metabolismo , Quebras de DNA de Cadeia Dupla , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Proteína do Grupo de Complementação A da Anemia de Fanconi/metabolismo , Proteína do Grupo de Complementação G da Anemia de Fanconi/metabolismo , Proteínas de Grupos de Complementação da Anemia de Fanconi/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Humanos , Mariposas , Osteoblastos/citologia , Osteoblastos/metabolismo , Proteína Rad52 de Recombinação e Reparo de DNA/genética , Proteína Rad52 de Recombinação e Reparo de DNA/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
4.
Nucleic Acids Res ; 51(17): 9166-9182, 2023 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-37503842

RESUMO

Histone deacetylase 6 (HDAC6) mediates DNA damage signaling by regulating the mismatch repair and nucleotide excision repair pathways. Whether HDAC6 also mediates DNA double-strand break (DSB) repair is unclear. Here, we report that HDAC6 negatively regulates DSB repair in an enzyme activity-independent manner. In unstressed cells, HDAC6 interacts with H2A/H2A.X to prevent its interaction with the E3 ligase RNF168. Upon sensing DSBs, RNF168 rapidly ubiquitinates HDAC6 at lysine 116, leading to HDAC6 proteasomal degradation and a restored interaction between RNF168 and H2A/H2A.X. H2A/H2A.X is ubiquitinated by RNF168, precipitating the recruitment of DSB repair factors (including 53BP1 and BRCA1) to chromatin and subsequent DNA repair. These findings reveal novel regulatory machinery based on an HDAC6-RNF168 axis that regulates the H2A/H2A.X ubiquitination status. Interfering with this axis might be leveraged to disrupt a key mechanism of cancer cell resistance to genotoxic damage and form a potential therapeutic strategy for cancer.


Assuntos
Reparo do DNA , Humanos , Linhagem Celular Tumoral , Dano ao DNA , Desacetilase 6 de Histona/genética , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação
5.
BMC Biol ; 22(1): 85, 2024 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-38627785

RESUMO

BACKGROUND: Inadequate DNA damage repair promotes aberrant differentiation of mammary epithelial cells. Mammary luminal cell fate is mainly determined by a few transcription factors including GATA3. We previously reported that GATA3 functions downstream of BRCA1 to suppress aberrant differentiation in breast cancer. How GATA3 impacts DNA damage repair preventing aberrant cell differentiation in breast cancer remains elusive. We previously demonstrated that loss of p18, a cell cycle inhibitor, in mice induces luminal-type mammary tumors, whereas depletion of either Brca1 or Gata3 in p18 null mice leads to basal-like breast cancers (BLBCs) with activation of epithelial-mesenchymal transition (EMT). We took advantage of these mutant mice to examine the role of Gata3 as well as the interaction of Gata3 and Brca1 in DNA damage repair in mammary tumorigenesis. RESULTS: Depletion of Gata3, like that of Brca1, promoted DNA damage accumulation in breast cancer cells in vitro and in basal-like breast cancers in vivo. Reconstitution of Gata3 improved DNA damage repair in Brca1-deficient mammary tumorigenesis. Overexpression of GATA3 promoted homologous recombination (HR)-mediated DNA damage repair and restored HR efficiency of BRCA1-deficient cells. Depletion of Gata3 sensitized tumor cells to PARP inhibitor (PARPi), and reconstitution of Gata3 enhanced resistance of Brca1-deficient tumor cells to PARP inhibitor. CONCLUSIONS: These results demonstrate that Gata3 functions downstream of BRCA1 to promote DNA damage repair and suppress dedifferentiation in mammary tumorigenesis and progression. Our findings suggest that PARP inhibitors are effective for the treatment of GATA3-deficient BLBCs.


Assuntos
Neoplasias Mamárias Animais , Inibidores de Poli(ADP-Ribose) Polimerases , Animais , Camundongos , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Dano ao DNA , Reparo do DNA , Neoplasias Mamárias Animais/genética , Neoplasias Mamárias Animais/patologia , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia
6.
J Biol Chem ; 299(3): 102975, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36738787

RESUMO

Ca2+ and voltage-activated K+ (BK) channels are ubiquitous ion channels that can be modulated by accessory proteins, including ß, γ, and LINGO1 BK subunits. In this study, we utilized a combination of site-directed mutagenesis, patch clamp electrophysiology, and molecular modeling to investigate if the biophysical properties of BK currents were affected by coexpression of LINGO2 and to examine how they are regulated by oxidation. We demonstrate that LINGO2 is a regulator of BK channels, since its coexpression with BK channels yields rapid inactivating currents, the activation of which is shifted ∼-30 mV compared to that of BKα currents. Furthermore, we show the oxidation of BK:LINGO2 currents (by exposure to epifluorescence illumination or chloramine-T) abolished inactivation. The effect of illumination depended on the presence of GFP, suggesting that it released free radicals which oxidized cysteine or methionine residues. In addition, the oxidation effects were resistant to treatment with the cysteine-specific reducing agent DTT, suggesting that methionine rather than cysteine residues may be involved. Our data with synthetic LINGO2 tail peptides further demonstrate that the rate of inactivation was slowed when residues M603 or M605 were oxidized, and practically abolished when both were oxidized. Taken together, these data demonstrate that both methionine residues in the LINGO2 tail mediate the effect of oxidation on BK:LINGO2 channels. Our molecular modeling suggests that methionine oxidation reduces the lipophilicity of the tail, thus preventing it from occluding the pore of the BK channel.


Assuntos
Cisteína , Canais de Potássio Ativados por Cálcio de Condutância Alta , Canais de Potássio Ativados por Cálcio de Condutância Alta/genética , Canais de Potássio Ativados por Cálcio de Condutância Alta/metabolismo , Cisteína/metabolismo , Oxirredução , Peptídeos/metabolismo , Metionina/metabolismo , Cálcio/metabolismo
7.
Anal Chem ; 96(21): 8586-8593, 2024 05 28.
Artigo em Inglês | MEDLINE | ID: mdl-38728058

RESUMO

Nowadays, signal enhancement is imperative to increase sensitivity of advanced ECL devices for expediting their promising applications in clinic. In this work, photodynamic-assisted electrochemiluminescence (PDECL) device was constructed for precision diagnosis of Parkinson, where an advanced emitter was prepared by electrostatically linking 2,6-dimethyl-8-(3-carboxyphenyl)4,4'-difluoroboradiazene (BET) with 1-butyl-3-methylimidazole tetrafluoroborate ([BMIm][BF4]). Specifically, protoporphyrin IX (PPIX) can trigger the photodynamic reaction under light irradiation with a wavelength of 450 nm to generate lots of singlet oxygen (1O2), showing a 2.43-fold magnification in the ECL responses. Then, the aptamer (Apt) was assembled on the functional BET-[BMIm] for constructing a "signal off" ECL biosensor. Later on, the PPIX was embedded into the G-quadruplex (G4) of the Apt to magnify the ECL signals for bioanalysis of α-synuclein (α-syn) under light excitation. In the optimized surroundings, the resulting PDECL sensor has a broad linear range of 100.0 aM ∼ 10.0 fM and a low limit of detection (LOD) of 63 aM, coupled by differentiating Parkinson patients from normal individuals according to the receiver operating characteristic (ROC) curve analysis of actual blood samples. Such research holds great promise for synthesis of other advanced luminophores, combined with achieving an early clinical diagnosis.


Assuntos
Compostos de Boro , Técnicas Eletroquímicas , Medições Luminescentes , Doença de Parkinson , Humanos , Doença de Parkinson/diagnóstico , Doença de Parkinson/sangue , Compostos de Boro/química , Técnicas Biossensoriais/métodos , alfa-Sinucleína/análise , alfa-Sinucleína/sangue , Protoporfirinas/química , Aptâmeros de Nucleotídeos/química , Limite de Detecção
8.
Analyst ; 149(2): 426-434, 2024 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-38099364

RESUMO

Nowadays, organic emitters suffer from insufficient electrochemiluminescence (ECL) efficiency in aqueous solutions, and their practical applications are severely restricted in the bio-sensing field. In this work, palladium nanospheres-embedded metal-organic frameworks (Pd@MOFs) were exploited to enhance the ECL efficiency of 2,6-dimethyl-8-(3-carboxyphenyl)4,4'-difluoroboradiazene (BET) prepared by a one-pot method in aqueous environment. First, the Pd@MOFs were generated via in situ reduction of Pd nanospheres anchored onto the MOFs, and fabricated by orderly coordination of palladium chloride (PdCl2) with 1,2,4,5-benzenetetramine (BTA) tetrahydrochloride. Then, the influence of protons on the ECL response of BET was studied in detail to obtain stronger ECL emission using potassium persulfate (K2S2O8) as co-reactant in aqueous environment. As a result, a 1.47-fold ECL efficiency enlargement of BET/K2S2O8 was harvested at the Pd@MOFs/GCE, where Ru(bpy)32+ behaved as a standard. Based on the fact that the ECL signals of the BET-covered Pd@MOFs modified glassy carbon electrode (simplified as BET/Pd@MOFs/GCE) can be quenched by Cu2+, the as-built ECL sensor showed a wide linear range (1.0-100.0 pM) and a limit of detection (LOD) as low as 0.12 pM. Hence, such research offers huge potential to promote the development of organic emitters in ECL biosensors and environmental monitoring.

9.
Proc Natl Acad Sci U S A ; 118(32)2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34353917

RESUMO

The increasing complexity of different cell types revealed by single-cell analysis of tissues presents challenges in efficiently elucidating their functions. Here we show, using prostate as a model tissue, that primary organoids and freshly isolated epithelial cells can be CRISPR edited ex vivo using Cas9-sgRNA (guide RNA) ribotnucleoprotein complex technology, then orthotopically transferred in vivo into immunocompetent or immunodeficient mice to generate cancer models with phenotypes resembling those seen in traditional genetically engineered mouse models. Large intrachromosomal (∼2 Mb) or multigenic deletions can be engineered efficiently without the need for selection, including in isolated subpopulations to address cell-of-origin questions.


Assuntos
Deleção Cromossômica , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Edição de Genes/métodos , Próstata/citologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteína 9 Associada à CRISPR/genética , Células Epiteliais , Genes Supressores de Tumor , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Organoides , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , RNA Guia de Cinetoplastídeos , Ribonucleoproteínas/genética , Regulador Transcricional ERG/genética , Ensaios Antitumorais Modelo de Xenoenxerto
10.
J Lipid Res ; 64(12): 100465, 2023 12.
Artigo em Inglês | MEDLINE | ID: mdl-37890669

RESUMO

Accurate intracellular cholesterol traffic plays crucial roles. Niemann Pick type C (NPC) proteins NPC1 and NPC2, are two lysosomal cholesterol transporters that mediate the cholesterol exit from lysosomes. However, other proteins involved in this process remain poorly defined. Here, we find that the previously unannotated protein TMEM241 is required for cholesterol egressing from lysosomes through amphotericin B-based genome-wide CRISPR-Cas9 KO screening. Ablation of TMEM241 caused impaired sorting of NPC2, a protein utilizes the mannose-6-phosphate (M6P) modification for lysosomal targeting, resulting in cholesterol accumulation in the lysosomes. TMEM241 is a member of solute transporters 35 nucleotide sugar transporters family and localizes on the cis-Golgi network. Our data indicate that TMEM241 transports UDP-N-acetylglucosamine (UDP-GlcNAc) into Golgi lumen and UDP-GlcNAc is used for the M6P modification of proteins including NPC2. Furthermore, Tmem241-deficient mice display cholesterol accumulation in pulmonary cells and behave pulmonary injury and hypokinesia. Taken together, we demonstrate that TMEM241 is a Golgi-localized UDP-GlcNAc transporter and loss of TMEM241 causes cholesterol accumulation in lysosomes because of the impaired M6P-dependent lysosomal targeting of NPC2.


Assuntos
Colesterol , Proteínas de Transporte Vesicular , Animais , Camundongos , Proteínas de Transporte Vesicular/metabolismo , Colesterol/metabolismo , Difosfato de Uridina/metabolismo , Lisossomos/metabolismo
11.
Anal Chem ; 95(50): 18572-18578, 2023 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-38064592

RESUMO

Electrochemiluminescence (ECL) has attracted significant interest in the analysis of cancer cells, where the ruthenium(II)-based emitter demonstrates urgency and feasibility to improve the ECL efficiency. In this work, the self-enhanced ECL luminophore was prepared by covalent anchoring of Pd nanoclusters on aminated metal organic frameworks (Pd NCs@MOFs), followed by linkage with bis(2,2'-bipyridine)-5-amino-1,10-phenanthroline ruthenium(II) (RuP). The resultant luminophore showed 214-fold self-magnification in the ECL efficiency over RuP alone, combined by promoting the interfacial photoelectron transfer. The enhanced mechanism through ion annihilation was critically proved by controlled experiments and density functional theory (DFT) calculations. Based on the above, a "signal off" ECL biosensor was built by assembly of tyrosine kinase 7 (PTK-7) aptamer (Apt) on the established sensing platform for analysis of human lung cancer cells (A549). The built sensor showed a lower detection limit of 8 cells mL-1, achieving the single-cell detection. This work reported a self-enhanced strategy for synthesis of advanced ECL emitters, combined by exploring the ECL biosensing devices in the single-cell analysis of cancers.


Assuntos
Técnicas Biossensoriais , Neoplasias Pulmonares , Nanopartículas Metálicas , Estruturas Metalorgânicas , Rutênio , Humanos , Medições Luminescentes , Técnicas Eletroquímicas , Limite de Detecção
12.
Anal Chem ; 95(10): 4735-4743, 2023 03 14.
Artigo em Inglês | MEDLINE | ID: mdl-36852949

RESUMO

Nowadays, electrochemiluminescence (ECL) efficiency of an organic emitter is closely related with its potential applications in food safety and environmental monitoring fields. In this work, 2,4,6-tris(4-carboxyphenyl)-1,3,5-triazine (TATB) was self-assembled to form hydrogen bond organic frameworks (HOFs), which worked as ideal reactors to generate highly active oxygen-containing radicals, followed by linking with isoluminol (ILu) via amide bond (termed ILu-HOFs). After covalent assembly with aminated indium-tin oxide electrode (labeled NH2-ITO), the ECL efficiency of the ILu-HOFs NH2-ITO showed about a 23.4-time increase over that of ILu itself in the presence of H2O2. Meanwhile, the enhanced ECL mechanism was mainly studied by electron paramagnetic resonance, theoretical calculation, and electrochemistry. On the above foundation, an aptamer "sandwich" ECL biosensor was constructed for detecting isocarbophos (ICP) via in situ elimination of H2O2 with catalase-linked palladium nanocubes (CAT-Pd NCs). The as-built sensor showed a broad linear range (1 pM to 100 nM) and a low limit of detection (LOD) down to 0.4 pM, coupled with efficient assays of ICP in lake water and cucumber juice samples. This strategy provides an effective way for the synthesis of advanced ECL emitter, coupled by showing promising applications in environmental and food analysis.


Assuntos
Técnicas Biossensoriais , Peróxido de Hidrogênio , Ligação de Hidrogênio , Medições Luminescentes , Limite de Detecção , Eletrodos , Técnicas Eletroquímicas
13.
J Virol ; 96(6): e0148021, 2022 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-35107379

RESUMO

In our previous study, we found that a new type of Chikungunya virus particle with a complete capsid deletion (ΔC-CHIKV) is still infectious in BHK-21 cells and demonstrated its potential as a live attenuated vaccine candidate. However, the low yield as well as the disability to propagate in vaccine production cell line Vero of ΔC-CHIKV are not practical for commercial vaccine development. In this study, we not only achieved the successful propagation of the viral particle in Vero cells, but significantly improved its yield through construction of a chimeric VEEV-ΔC-CHIKV and extensive passage in Vero cells. Mechanistically, high production of VEEV-ΔC-CHIKV is due to the improvement of viral RNA packaging efficiency conferred by adaptive mutations, especially those in envelope proteins. Similar to ΔC-CHIKV, the passaged VEEV-ΔC-CHIKV is safe, immunogenic, and efficacious, which protects mice from CHIKV challenge after only one shot of immunization. Our study demonstrates that the utilization of infectious capsidless viral particle of CHIKV as a vaccine candidate is a practical strategy for the development of alphavirus vaccine. IMPORTANCE Chikungunya virus (CHIKV) is one of important emerging alphaviruses. Currently, there are no licensed vaccines against CHIKV infection. We have previously found a new type of Chikungunya virus particle with a complete capsid deletion (ΔC-CHIKV) as a live attenuated vaccine candidate that is not suitable for commercial vaccine development with the low viral titer production. In this study, we significantly improved its production through construction of a chimeric VEEV-ΔC-CHIKV. Our results proved the utilization of infectious capsidless viral particle of CHIKV as a safe and practical vaccine candidate.


Assuntos
Febre de Chikungunya , Vírus Chikungunya , Vacinas Virais , Cultura de Vírus , Animais , Proteínas do Capsídeo/genética , Febre de Chikungunya/prevenção & controle , Vírus Chikungunya/genética , Chlorocebus aethiops , Camundongos , Desenvolvimento de Vacinas , Vacinas Atenuadas/genética , Células Vero , Vacinas Virais/genética , Cultura de Vírus/métodos
14.
Appl Opt ; 62(36): 9512-9522, 2023 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-38108776

RESUMO

In this paper, we present a robust method for nonisotropic point light source calibration through feature points selection. By analyzing the relationship between the observed surface and its image intensity under near-field lighting, the feature points selection method is first developed to effectively address the noisy observations and improve calibration robustness. Afterward, to enhance efficiency and accuracy of the calibration, a cost function of l p-norm is established based on the above relationship, and an improved Newton method-based iteration process is applied to calculate the light source parameters. The simulations demonstrate that the proposed method is capable of achieving robust calibration results with the estimation error less than 2.7 mm and 0.8°, even though the image intensities are corrupted by Gaussian white noise with standard deviation up to 0.4. The experimental validation is performed using a self-designed photometric stereo system, where the calibration of point light sources is conducted, and measurements are taken on a standard sphere and compressor blade based on the obtained calibration results, which demonstrates the effectiveness of what we believe to be a new method.

15.
Pestic Biochem Physiol ; 193: 105446, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37248015

RESUMO

The use of herbicides is believed to have an impact on the metabolism, physiology and biochemistry of fish. In this study, we studied the effects of metamifop on the production and metabolism of Monopterus. albus living in the water. According to the semi-lethal concentration of metamifop for 96 h, four MET concentration groups (0.2-, 0.4-, 0.6- and 0.8 mg L-1) were set up for 96 h exposure test. The ammonia discharge rate decreased, hemolymph ammonia content increased significantly, and hemolymph urea nitrogen content decreased at all time periods of metamifop exposure. In liver, the protein content decreased, the neutral protease content increased significantly (p < 0.01), amino acid content increased, and ATP content increased significantly (p < 0.01). In brain, the protein content increased, the activity of acid protease, neutral protease and alkaline protease all decreased, amino acid content decreased significantly (p < 0.01), and the content of ATP decreased. Glutamic-pyruvic transaminase (GPT) activity did not change in liver but decreased in brain. Glutamine synthetase (GS) activity decreased in liver and increased in brain. Glutaminase (GLS) activity decreased in liver and increased in brain. In conclusion, the liver and brain tissues of M. albus react differently to MET exposure. The liver mainly synthesizes energy through hydrolyzed protein, while the brain mainly synthesizes protein. Amino acids produced by protein hydrolysis cannot be converted to alanine for storage, and the degraded amino acids lead to the elevation of endogenous ammonia. MET inhibits the removal of ammonia from M. albus. Only liver tissue can detoxify the eel by converting ammonia into glutamine. Brain should have to tolerate high levels of endogenous ammonia.


Assuntos
Amônia , Smegmamorpha , Animais , Amônia/metabolismo , Aminoácidos/metabolismo , Glutamina/metabolismo , Fígado/metabolismo , Smegmamorpha/metabolismo , Trifosfato de Adenosina/metabolismo , Glutamato-Amônia Ligase/metabolismo , Ureia/metabolismo
16.
Anal Chem ; 94(8): 3708-3717, 2022 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-35172575

RESUMO

Nowadays, aggregation quenching of most organic photosensitizers in aqueous media seriously restricts analytical and biomedical applications of photoelectrochemical (PEC) sensors. In this work, an aggregation-enhanced PEC photosensitizer was prepared by electrostatically bonding protoporphyrin IX (PPIX) with an ionic liquid of 1-butyl-3-methylimidazole tetrafluoroborate ([BMIm][BF4]), termed as PPIX-[BMIm] for clarity. The resultant PPIX-[BMIm] showed weak photocurrent in pure dimethyl sulfoxide (DMSO, good solvent), while the PEC signals displayed a 44.1-fold enhancement in a water (poor solvent)/DMSO binary solvent with a water fraction (fw) of 90%. Such PEC-enhanced mechanism was critically studied by electrochemistry and density functional theory (DFT) calculation in some detail. Afterward, a label-free PEC cytosensor was built for ultrasensitive bioassay of acute lymphoblastic leukemia (molt-4) cells by electrodepositing Au nanoparticles (Au NPs) on the PPIX-[BMIm] aggregates and sequential assembly of protein tyrosine kinase (PTK) aptamer DNA (aptDNA). The resultant cytosensor showed a wide linear range (300 to 3 × 105 cells mL-1) with a limit of detection (LOD) as low as 63 cells mL-1. The aggregation-enhanced PEC performance offers a valuable and practical pathway for synthesis of advanced organic photosensitizer to explore its PEC applications in early diagnosis of tumors.


Assuntos
Técnicas Biossensoriais , Líquidos Iônicos , Nanopartículas Metálicas , Técnicas Eletroquímicas , Ouro , Fármacos Fotossensibilizantes , Protoporfirinas , Eletricidade Estática
17.
Opt Express ; 30(7): 11912-11922, 2022 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-35473124

RESUMO

White light scanning interferometry (WLSI) has been an extremely powerful technique in precision measurements. In this work, a phase noise estimation based surface recovery algorithm is proposed, which can significantly improve the measurement accuracy by decreasing the noise level in phase map coming from the systemic and environmental disturbances. The noise existed in phase map is firstly researched in spectrum domain and defined as the linear combination of complex terms at each angular wavenumber. Afterwards, based on the theoretical linearity of the phase distribution, the surface features can be redefined through establishing the function with respect to phase noise. By applying least square estimation (LSE), a spectral coefficient is defined to determine the optimal estimation of phase noise that represents the best statistical consistency with the actual case, from which a more accurate surface after removing most phase noise will then be generated. In order to testify the noise elimination ability of the proposed method, a nano-scale step height standard (9.5nm±1.0nm) is scanned, and the measurement result 9.49nm with repeatability 0.17nm is successfully achieved. Moreover, a leading edge of an aero-engine blade is also tested to investigate the potential of this method in industrial inspections. The measurement comparison with AFM is also displayed.

18.
Microb Pathog ; 163: 105387, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34990781

RESUMO

The toxicity of polystyrene nano/microplastics with diameter sizes of 50um and 100 nm and concentrations of 100 and 1000 µg/mL on gut microbiota, antioxidant activity and innate immune response in zebrafish was investigated. After exposure to polystyrene plastics particle, the pathological morphological changes of intestine and gills were observed, and the injury severity was related to the concentration and particle size of plastics. Significant changes in the richness and diversity of gut microbiota were observed after polystyrene plastics-exposed in zebrafish. The plastics-treated groups exhibited more substantial oxidative stress than the control group. In addition, the mRNA expression level of most pro- and anti-inflammatory factors, including IL-8, NF-κb, and IL-10, increased while the mRNA expression of TNF-α, a pro-inflammatory factor, decreased. Our results suggest that polystyrene nano/microplastics may represent a potential threat to the gut microbiota, oxidative status, and innate immunity. These results indicated that polystyrene nano/microplastics exerted size and concentration-dependent toxicity on zebrafish. The findings provide new evidence for the toxicity of polystyrene plastics on zebrafish.


Assuntos
Microbiota , Poluentes Químicos da Água , Animais , Disbiose/induzido quimicamente , Imunidade Inata , Microplásticos , Estresse Oxidativo , Plásticos , Poliestirenos , Poluentes Químicos da Água/análise , Peixe-Zebra
19.
Rev Cardiovasc Med ; 23(10): 333, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-39077142

RESUMO

Background: Neutrophil percentage to albumin ratio (NPAR) has been shown to be correlated with the prognosis of various diseases. This study aimed to explore the effect of NPAR on the prognosis of patients in coronary care units (CCU). Method: All data in this study were extracted from the Medical Information Mart for Intensive Care III (MIMIC-III, version1.4) database. All patients were divided into four groups according to their NPAR quartiles. The primary outcome was in-hospital mortality. Secondary outcomes were 30-day mortality, 365-day mortality, length of CCU stay, length of hospital stay, acute kidney injury (AKI), and continuous renal replacement therapy (CRRT). A multivariate binary logistic regression analysis was performed to confirm the independent effects of NPAR. Cox regression analysis was performed to analyze the association between NPAR and 365-day mortality. The curve in line with overall trend was drawn by local weighted regression (Lowess). Subgroup analysis was used to determine the effect of NPAR on in-hospital mortality in different subgroups. Receiver operating characteristic (ROC) curves were used to evaluate the ability of NPAR to predict in-hospital mortality. Kaplan-Meier curves were constructed to compare the cumulative survival rates among different groups. Result: A total of 2364 patients in CCU were enrolled in this study. The in-hospital mortality rate increased significantly as the NPAR quartiles increased (p < 0.001). In multivariate logistic regression analysis, NPAR was independently associated with in-hospital mortality (quartile 4 versus quartile 1: odds ratio [OR], 95% confidence interval [CI]: 1.83, 1.20-2.79, p = 0.005, p for trend < 0.001). In Cox regression analysis, NPAR was independently associated with 365-day mortality (quartile 4 versus quartile 1: OR, 95% CI: 1.62, 1.16-2.28, p = 0.005, p for trend < 0.001). The Lowess curves showed a positive relationship between NPAR and in-hospital mortality. The moderate ability of NPAR to predict in-hospital mortality was demonstrated through ROC curves. The area under the curves (AUC) of NPAR was 0.653 (p < 0.001), which is better than that of the platelet to lymphocyte ratio (PLR) (p < 0.001) and neutrophil count (p < 0.001) but lower than the Sequential Organ Failure Assessment (p = 0.046) and Simplified Acute Physiology Score II (p < 0.001). Subgroup analysis did not reveal any obvious interactions in most subgroups. However, Kaplan-Meier curves showed that as NPAR quartiles increased, the 30-day (log-rank, p < 0.001) and 365-day (log-rank, p < 0.001) cumulative survival rates decreased significantly. NPAR was also independently associated with AKI (quartile 4 versus quartile 1: OR, 95% CI: 1.57, 1.19-2.07, p = 0.002, p for trend = 0.001). The CCU and hospital stay length was significantly prolonged in the higher NPAR quartiles. Conclusions: NPAR is an independent risk factor for in-hospital mortality in patients in CCU and has a moderate ability to predict in-hospital mortality.

20.
Org Biomol Chem ; 20(27): 5452-5462, 2022 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-35770913

RESUMO

We have developed an improved cyanide-free strategy for the synthesis of glycosyl carboxylic acids, employing stereoselective C-vinyl glycosylation and oxidative cleavage of C-vinyl glycosides as key steps. Compared to our previous work, the amount of NaIO4 required for the oxidative cleavage step is reduced significantly from 18 equivalents to 4.5 equivalents. This modification not only is advantageous in terms of operation and costs but also avoids the over-oxidation problem, thus greatly expanding the substrate scope, which is evidenced by the fact that 10 out of 21 glycosyl carboxylic acids synthesized are undocumented. With differently O5-protected furanosyl acids in hand, we demonstrate that an electron-rich protecting group is beneficial for the decarboxylative arylation of furanosyl carboxylic acids. This represents a rare example of protecting groups affecting the reaction efficiency in radical C-glycosylation. As C-vinyl glycosides can be prepared stereoselectively and the oxidative step is stereoretentive, the approach provides an effective means to access 1,2-trans or 1,2-cis glycosyl acids, which would be a valuable alternative to the cyanide-based synthesis of glycosyl carboxylic acids.


Assuntos
Ácidos Carboxílicos , Glicosídeos , Glicosilação , Estresse Oxidativo , Estereoisomerismo
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