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Selective and sensitive imaging of intracellular mature microRNAs (miRNAs) is of great importance for biological process study and medical diagnostics. However, this goal remains challenging because of the interference of precursor miRNAs (pre-miRNAs) and the low abundance of mature miRNAs. Herein, we develop an endogenous enzyme-driven amplified DNA nanocage probe (Acage) for the selective and sensitive imaging of mature miRNAs in living cells. The Acage consists of a microRNA-responsive probe, an endogenous enzyme-driven fuel strand, and a DNA nanocage framework with an inner cavity. Benefiting from the size selectivity of DNA nanocage, smaller mature miRNAs rather than larger pre-miRNAs are allowed to enter the cavity of DNA nanocage for molecular recognition; thus, Acage can significantly reduce the signal interference of pre-miRNAs. Moreover, with the driving force of an endogenous enzyme apurinic/apyrimidinic endonuclease 1 (APE1) for efficient signal amplification, Acage enables sensitive intracellular miRNA imaging without an additional external intervention. With these features, Acage was successfully applied for intracellular imaging of mature miRNAs during drug treatment. We believe that this strategy provides a promising pathway for better understanding the functions of mature microRNAs in biological processes and medical diagnostics.
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Sondas de ADN , MicroARNs , MicroARNs/análisis , MicroARNs/metabolismo , Humanos , Sondas de ADN/química , Nanoestructuras/química , Imagen Óptica , Células HeLaRESUMEN
Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas12a nucleases have emerged as a promising alternative to CRISPR-Cas9 in gene editing and expression regulation. However, the adoption of Cas12a has been hindered due to general off-target activities and limited efficiency. Here, we utilized a hybrid engineered Cas12a variant and hairpin-spacer crRNAs (h-CAP) to enhance the specificity and efficiency of the CRISPR-Cas12a system. Leveraging the h-CAP strategy, we demonstrate both single-base-specific and multiplex gene expression regulation in human cells.
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Sistemas CRISPR-Cas , Edición Génica , Humanos , Sistemas CRISPR-Cas/genética , Endonucleasas/metabolismoRESUMEN
Lead is a widespread environmental hazard that can adversely affect multiple biological functions. Blood cells are the initial targets that face lead exposure. However, a systematic assessment of lead dynamics in blood cells at single-cell resolution is still absent. Herein, C57BL/6 mice were fed with lead-contaminated food. Peripheral blood was harvested at different days. Extracted red blood cells and leukocytes were stained with 19 metal-conjugated antibodies and analyzed by mass cytometry. We quantified the time-lapse lead levels in 12 major blood cell subpopulations and established the distribution of lead heterogeneity. Our results show that the lead levels in all major blood cell subtypes follow lognormal distributions but with distinctively individual skewness. The lognormal distribution suggests a multiplicative accumulation of lead with stochastic turnover of cells, which allows us to estimate the lead lifespan of different blood cell populations by calculating the distribution skewness. These findings suggest that lead accumulation by single blood cells follows a stochastic multiplicative process.
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Plomo , Longevidad , Animales , Ratones , Plomo/toxicidad , Ratones Endogámicos C57BL , Leucocitos , EritrocitosRESUMEN
BACKGROUND: Endogenous endophthalmitis (EE) is a rare but highly destructive eye emergency secondary to systemic infection. Acute endophthalmitis can lead to irreversible vision impairment or even loss of the whole eye, unless being diagnosed and treated promptly. CASE PRESENTATION: This study reports three typical EE cases of endogenous endophthalmitis secondary to different severe systemic diseases. Patients were recruited from the Department of ophthalmology at Zhongnan hospital of Wuhan University and the Department of ophthalmology at the Second Affiliated Hospital of Fujian Medical University. Patients were followed up for up to 60 days. Among these cases, the eye symptoms is the initial manifestations while secondary to original different special systemic conditions. Patients have been treated under dynamically prompt response undergoing systemic treatment and eye treatment at the same time. Best corrected visual acuity were 20/40, 20/60 and light perception during follow-up evaluation. CONCLUSIONS: Our observation suggest that prompt identification and treatment could save patients' vision from EE.
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Endoftalmitis , Infecciones Bacterianas del Ojo , Agudeza Visual , Humanos , Antibacterianos/uso terapéutico , Endoftalmitis/diagnóstico , Endoftalmitis/microbiología , Infecciones Bacterianas del Ojo/microbiología , Infecciones Bacterianas del Ojo/diagnóstico , Infecciones Bacterianas del Ojo/tratamiento farmacológico , Agudeza Visual/fisiologíaRESUMEN
Clubroot, caused by Plasmodiophora brassicae, is a globally destructive soil-borne disease affecting cruciferous plants. Here, the predominant pathotypes of P. brassicae in six cities within Zhejiang Province were identified using the Williams and European Clubroot Differential (ECD) systems. A phylogenetic analysis of P. brassicae isolates infecting cruciferous crops worldwide was conducted using MEGA, and their ITS2 secondary structures were predicted through the ITS2 database. Accessions of B. rapa, B. oleracea, B. juncea, and Eruca sativa Mill. were employed to assess clubroot resistance. The results revealed that the prevalent pathotypes in Zhejiang Province were pathotype 1, ECD20/31/12 and ECD24/16/30; pathotype 3, ECD20/15/4; pathotype 8, ECD16/0/0 and ECD24/0/0; and pathotype 2, ECD16/15/15. Isolates from distinct genera of Brassicaceae formed separate branches in the evolutionary tree. Moreover, isolates of Brassica crops from Zhejiang Province exhibited homology with those from other global regions, a finding corroborated by their ITS2 secondary structure. Approximately 80% and 95% of B. rapa and B. juncea crops displayed susceptible phenotypes for pathotype 8, ECD16/0/0, whereas approximately 60% of B. oleracea crops exhibited resistance. Furthermore, three Brassica crop accessions showed significant variation in resistance to the pathogen, both among morphological and geographical origin groups. This study contributes to understanding the distribution of diverse P. brassicae pathotypes in different regions of Zhejiang Province and facilitates the identification of Brassica crops with potential disease resistance suitable for cultivation in the province.
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BACKGROUND: Antarctic krill peptide (AKP) has gained considerable interest because of its multiple biological functions. However, its application may be limited by its poor stability and susceptibility to degradation. Encapsulation of AKP using a nanoparticle delivery system is an effective way to overcome these problems. In the present study, bovine serum albumin (BSA) and chitosan (CS) were used as delivery vehicles to encapsulate AKP. RESULTS: The results revealed that the particle size (83.3 ± 4.4-222.4 ± 32.7 nm) and zeta-potential (35.1 ± 0.7-45.0 ± 2.7 mV) of nanoparticles (NPs) increased with the increasing content of BSA, but the polydispersity index decreased (1.000 ± 0.002 to 0.306 ± 0.011). Hydrogen bonding, hydrophobic and electrostatic interactions were the main forces to form BSA/CS-AKP NPs. X-ray diffraction revealed that AKP was encapsulated by BSA/CS. Scanning electron microscopy images exhibited that the NPs were spherical in shape, uniform in size and tightly bound. BSA/CS-AKP NPs exhibited excellent stability in the pH range (2-5) and after 15 days of storage, and could hinder the release of AKP in simulated gastric environment and promote the release of AKP in simulated intestinal environment. After simulated digestion, the hypoglycemic activity of encapsulated AKP was better than that of unencapsulated AKP. CONCLUSION: Our results revealed that the BSA/CS showed great potential for protecting and delivering AKP. © 2024 Society of Chemical Industry.
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Emerging studies have reported the expansion of myeloid-derived suppressor cells (MDSCs) in some autoimmune disorders, such as systemic lupus erythematosus (SLE), but the detailed molecular mechanisms of the aberrant expansion in SLE are still unclear. In the present study, we confirmed that the increased MDSCs positively correlated with disease activity in SLE patients. The suppressive capacity of MDSCs from patients with high activity was lower than that of MDSCs from patients with low activity. Moreover, the potential precursors for MDSCs, common myeloid progenitors (CMPs) and granulocyte-monocyte progenitors (GMPs), were markedly increased in the bone marrow (BM) aspirates of SLE patients. As an important regulator of cell fate decisions, aberrant activation of Notch signalling was reported to participate in the pathogenesis of SLE. We found that the expression of Notch1 and its downstream target gene hairy and enhancer of split 1 (Hes-1) increased markedly in GMPs from SLE patients. Moreover, the Notch1 signalling inhibitor DAPT profoundly relieved disease progression and decreased the proportion of MDSCs in pristane-induced lupus mice. The frequency of GMPs was also decreased significantly in lupus mice after DAPT treatment. Furthermore, the inhibition of Notch1 signalling could limit the differentiation of MDSCs in vitro. The therapeutic effect of DAPT was also verified in Toll-like receptor 7 (TLR7) agonist-induced lupus mice. Taken together, our results demonstrated that Notch1 signalling played a crucial role in MDSC differentiation in SLE. These findings will provide a promising therapy for the treatment of SLE.
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Lupus Eritematoso Sistémico , Células Supresoras de Origen Mieloide , Animales , Ratones , Inhibidores de Agregación Plaquetaria/metabolismo , Inhibidores de Agregación Plaquetaria/farmacología , Diferenciación CelularRESUMEN
Open-heart surgery is associated with high morbidity, with acute kidney injury (AKI) being one of the most commonly observed postoperative complications. Following open-heart surgery, in an observational study we found significantly higher numbers of blood neutrophils in a group of 13 patients with AKI compared to 25 patients without AKI (AKI: 12.9±5.4 ×109 cells/L; non-AKI: 10.1±2. 9 ×109 cells/L). Elevated serum levels of neutrophil extracellular trap (NETs) components, such as dsDNA, histone 3, and DNA binding protein Y-box protein (YB)-1, were found within the first 24 hours in patients who later developed AKI. We could demonstrate that NET formation and hypoxia triggered the release of YB-1, which was subsequently shown to act as a mediator of kidney tubular damage. Experimentally, in two models of AKI mimicking kidney hypoperfusion during cardiac surgery (bilateral ischemia/reperfusion (I/R) and systemic lipopolysaccharide (LPS) administration), a neutralizing YB-1 antibody was administered to mice. In both models, prophylactic YB-1 antibody administration significantly reduced the tubular damage (damage score range 1-4, the LPS model: non-specific IgG control, 0.92±0.23; anti-YB-1 0.65±0.18; and in the I/R model: non-specific IgG control 2.42±0.23; anti-YB-1 1.86±0.44). Even in a therapeutic, delayed treatment model, antagonism of YB-1 ameliorated AKI (damage score, non-specific IgG control 3.03±0.31; anti-YB-1 2.58±0.18). Thus, blocking extracellular YB-1 reduced the effects induced by hypoxia and NET formation in the kidney and significantly limited AKI, suggesting that YB-1 is part of the NET formation process and an integral mediator of cross-organ effects.
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Lesión Renal Aguda , Trampas Extracelulares , Daño por Reperfusión , Ratones , Animales , Proteínas de Unión al ADN , Lipopolisacáridos , Riñón , Isquemia/complicaciones , Hipoxia , Inmunoglobulina G , Daño por Reperfusión/complicaciones , Ratones Endogámicos C57BLRESUMEN
The accurate, rapid, and sensitive identification of cancer cells in complex physiological environments is significant in biological studies, personalized medicine, and biomedical engineering. Inspired by the naturally confined enzymes on fluid cell membranes, a fluidly confined CRISPR-based DNA reporter (FINDER) was developed on living cell membranes, which was successfully applied for rapid and sensitive cancer cell identification in clinical blood samples. Benefiting from the spatial confinement effect for improved local concentration, and membrane fluidity for higher collision efficiency, the activity of CRISPR-Cas12a was, for the first time, found to be significantly enhanced on living cell membranes. This new phenomenon was then combined with multiple aptamer-based DNA logic gate for cell recognition, thus a FINDER system capable of accurate, rapid and sensitive cancer cell identification was constructed. The FINDER rapidly identified target cells in only 20â min, and achieved over 80 % recognition efficiency with only 0.1 % of target cells presented in clinical blood samples, indicating its potential application in biological studies, personalized medicine, and biomedical engineering.
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Técnicas Biosensibles , Neoplasias , Membrana Celular , ADN , Fluidez de la Membrana , Oligonucleótidos , Bioingeniería , Sistemas CRISPR-Cas/genética , Neoplasias/genéticaRESUMEN
BACKGROUND: Hormones play an indispensable role during fruit ripening, nine clades in 2-oxoglutarate-dependent dioxygenase (2OGD) superfamily are responsible for the hormone biosynthesis and metabolism, but less information is known about them. RESULTS: A total of 163 Vv2OGD superfamily members were identified from grape genome, which were mainly expanded by local (tandem and proximal) duplication. Phylogenetic analysis of 2OGD members in grape and Arabidopsis indicates 37 members in Vv2OGD superfamily are related to hormone biosynthesis and metabolism process (Vv2OGD-H), which could be divided into 9 clades, gibberellin (GA) 3-oxidase (GA3ox), GA 20-oxidase (GA20ox), carbon-19 GA 2-oxidase (C19-GA2ox), carbon-20 GA 2-oxidase (C20-GA2ox), 1-aminocyclopropane-1-carboxylic acid oxidase (ACO), dioxygenase for auxin oxidation (DAO), lateral branching oxidoreductas (LBO), downy mildew resistant 6 and DMR6-like oxygenase (DMR6/DLO) and jasmonate-induced oxygenase (JOX). Sixteen of these 37 Vv2OGD-Hs are expressed in grape berry, in which the expression patterns of VvGA2oxs, VvDAOs and VvJOXs shows a correlation with the change patterns of GAs, indole-3-acetic acid (IAA) and jasmonates (JAs), indicating the involvement of these genes in grape berry development by regulating corresponding hormones. Twelve Vv2OGD-Hs respond to methyl JA (MeJA) treatment, of which eight may lead to the inhibition of the ripening process by the crosstalk of JAs-salicylic acids (SAs), JAs-GAs and JAs-JAs, while seven Vv2OGD-Hs respond to ABA treatment may be responsible for the promotion of ripening process by the interplay of abscisic acid (ABA)-strigolactones (SLs), ABA-SAs, ABA-GAs, ABA-JAs. Especially, VvLBO1 reach an expression peak near véraison and up-regulate about four times after ABA treatment, which implies SLs and ABA-SLs crosstalk may be related to the onset of berry ripening in grape. CONCLUSIONS: This study provides valuable clues and new insights for the mechanism research of Vv2OGD-Hs in hormones regulation during the grape berry development.
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Arabidopsis , Dioxigenasas , Vitis , Ácido Abscísico/metabolismo , Arabidopsis/metabolismo , Dioxigenasas/genética , Dioxigenasas/metabolismo , Frutas , Regulación de la Expresión Génica de las Plantas , Hormonas/metabolismo , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Vitis/metabolismoRESUMEN
Cyclin-dependent kinases 4/6 (CDK4/6) and D1-type cyclins (CCND1) can regulate the pro-inflammatory functions of various cytokines during the inflammatory response. This study investigated the association between CDK4/6-CCND1 variants and susceptibility in patients with Behcet's disease (BD). This case-control study enrolled 542 patients with BD and 754 healthy controls. Fourteen tagged single nucleotide polymorphisms (tag SNPs) of the CDK4/6-CCND1 gene were genotyped using the Sequenom MassARRAY system and iPLEX® Pro assay. The results indicated that the frequency of the CDK6 rs2282983 TT genotype was higher in the BD group than the control group (Pc = 0.040, OR = 1.408, 95% CI = 1.124-1.765), and CDK6 rs2282983 CT and rs42034 AG were negatively associated with BD (Pc = 3.647 × 10-4, OR = 0.598, 95% CI = 0.471-0.758; Pc = 0.039, OR = 0.626, 95% CI = 0.459-0.852, respectively). Furthermore, statistical analysis showed that CDK6 rs2282983 TT and CT genotypes were significantly associated with skin lesions in patients with BD (Pc = 0.042, OR = 1.436, 95% CI = 1.130-1.824; Pc = 0.001, OR = 0.594, 95% CI = 0.461-0.764, respectively). This study suggests that the CDK6 loci rs2282983 and rs42034 might confer genetic susceptibility to BD in a Han Chinese population, which could provide new insights into the pathogenesis of BD.
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Síndrome de Behçet , Síndrome de Behçet/genética , Estudios de Casos y Controles , China/epidemiología , Quinasa 6 Dependiente de la Ciclina/genética , Quinasas Ciclina-Dependientes/genética , Ciclinas/genética , Citocinas/genética , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Polimorfismo de Nucleótido SimpleRESUMEN
BACKGROUND: The role of HOX transcript antisense RNA (HOTAIR) has been proven to be important in tumorigenesis. However, how this molecule promotes metastasis and invasion in PCa is still unclear. METHODS: The relationship between HOTAIR and hepatocellular adhesion molecule (hepaCAM) in PCa was identified by immunohistochemistry, immunofluorescence, plasmid transfection, quantitative real-time PCR and immunoblotting. The regulatory effects of HOTAIR on hepaCAM and MAPK signalling and their key roles in PCa metastasis were investigated in vitro. RESULTS: The expression of HOTAIR was inversely correlated with hepaCAM in the blood and tissue of PCa patients. Here, hepaCAM was identified as a novel target gene of HOTAIR and was critical for the invasiveness of PCa. HOTAIR recruited PRC2 to the hepaCAM promoter, resulting in high levels of H3K27me3 and the absence of hepaCAM with an abnormally activated MAPK pathway. Both HOTAIR depletion and EZH2 inhibition could induce hepaCAM re-expression with inhibitory MAPK signalling and decrease the invasive and metastatic capabilities of PCa cells. CONCLUSIONS: This study demonstrates that HOTAIR promotes invasion and metastasis of PCa by decreasing the inhibitory effect of hepaCAM on MAPK signalling. Therefore, the HOTAIR/hepaCAM/MAPK axis may provide a new avenue towards therapeutic strategies and prognostic indicators for advanced prostate cancer.
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Proteínas de Ciclo Celular/metabolismo , Regulación Neoplásica de la Expresión Génica/fisiología , Invasividad Neoplásica/genética , Neoplasias de la Próstata/patología , ARN Largo no Codificante/metabolismo , Anciano , Humanos , Masculino , Persona de Mediana Edad , Invasividad Neoplásica/patología , Neoplasias de la Próstata/genéticaRESUMEN
OBJECTIVES: MicroRNA-199b-3p (miR-199b-3p) plays a crucial role in the malignant development of various cancers, but little known in prostate cancer (PCa). The aim of our study was to demonstrate the function of miR-199b-3p in PCa. METHODS: Quantitative real-time polymerase chain reaction (RT-qPCR) was used to detect miR-199b-3p expression in PCa and benign prostatic hyperplasia (BPH) tissue samples. In addition, we examined the relationship between the poor prognosis in PCa and miR-199b-3p. Western blot was used to analyze the expression of Phospholipase Cε (PLCε). CCK8 and colony-forming assays were applied to detect the proliferation of PCa. EdU assay is used to detect PCa cells uptake of EdU. Luciferase reporter assay was applied to analyze the binding between miR-199b-3p and PLCε. RESULTS: It has been shown that miR-199b-3p in PCa was significantly lower than that in benign prostatic hyperplasia and correlated with poor prognosis. Meanwhile, upregulation of miR-199b-3p can prominently inhibit the proliferation of PCa cells, while its down-regulation triggered opposite result. PLCε was identified as the downstream binding target gene and negatively associated with that of miR-199b-3p. CONCLUSION: miR-199b-3p suppresses malignant proliferation by inhibiting PLCε in prostate cancer in vitro and vivo.
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MicroARNs/genética , Fosfoinositido Fosfolipasa C/antagonistas & inhibidores , Neoplasias de la Próstata/metabolismo , Animales , Línea Celular Tumoral , Movimiento Celular/fisiología , Proliferación Celular/fisiología , Regulación hacia Abajo , Xenoinjertos , Humanos , Masculino , Ratones , Ratones Desnudos , MicroARNs/metabolismo , Persona de Mediana Edad , Fosfoinositido Fosfolipasa C/genética , Fosfoinositido Fosfolipasa C/metabolismo , Pronóstico , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patologíaRESUMEN
MAIN CONCLUSION: Genome-wide identification, analysis and functional characterization of an unreported VvBBX gene showed a response to light and positive correlation with anthocyanin content, but also inhibition of light-induced anthocyanin synthesis. B-box (BBX) proteins are a class of zinc (Zn) finger transcription factors or regulators characterized by the presence of one or two BBX domains and play important roles in plant growth and development. However, the BBX genes' potential functions are insufficiently characterized in grape, a globally popular berry with high economic value. Here, 25 BBX family genes including a novel member (assigned VvBBX44) were identified genome widely in grape. The expression level of these VvBBXs were analyzed in 'Cabernet Sauvignon' (V. vinifera) stem, flower, leaf, tendril, petiole, and developing berries. The expression of VvBBX44 increased in developing 'Cabernet Sauvignon' berries. Its expression was inhibited in 'Jingxiu' and 'Muscat Hamburg' berry skin without sunlight. Furthermore, overexpression of VvBBX44 decreased the expression of LONG HYPOCOTYL 5 (VvHY5) and UDP-glucose flavonoid 3-O-glucosyltransferase (VvUFGT), and reduced the anthocyanin content in grape calli. Our results suggest that VvBBX44 may play an important role in grape berry coloring by directly repressing VvHY5 expression. This study provides new insights into the potential role of VvBBXs in berry development and light response and contributes to the understanding on the regulation mechanism of VvBBX44 in anthocyanin biosynthesis.
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Vitis , Antocianinas , Frutas/genética , Frutas/metabolismo , Regulación de la Expresión Génica de las Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Vitis/genética , Vitis/metabolismoRESUMEN
Aconitine linoleate (11) isolated from the Aconitum sinchiangense W. T. Wang exhibited significant anti-tumor activity. Based on this, a series of novel lipo-diterpenoid alkaloids were synthesized and evaluated for their anticancer activities against MCF-7 and MCF-7/ADR cell lines. Seventeen compounds, including 18-20, 22, 24-32, 36, 39, 41-42 possessed higher anti-proliferative activities (IC50 < 20 µM) against MCF-7 cell lines, which were better than the reference drug etoposide (IC50 = 18.01 ± 1.64 µM), among which compound 24 (IC50 = 4.00 ± 0.30 µM) was found to be the most potent derivative, being 4.5-fold more active than etoposide. Meanwhile, eighteen compounds, including 18-22, 24, 26-32, 36, 38-39, 41-42 presented excellent activities (IC50 < 20 µM) against MCF-7/ADR cell lines, better than etoposide (IC50 = 35.48 ± 0.29 µM) and doxorubicin (IC50 = 67.61 ± 6.5 µM). The most potent compound (19) was 13.5- and 25.7-fold more active than etoposide and doxorubicin against MCF-7/ADR cell lines, respectively. The structure-activity relationship (SAR) studies indicated that the 3-OH, 8-lipo, 14-benzene ring, and nitrogen atom with proper alkaline are crucial elements for anti-proliferative activity of target lipo-diterpenoid compounds. The proper length, the double bonds or di-fluoro-substituted at C-8 fatty acid chain, the para-donating electron group on 14-benzene group, and 13-OH are all favorable for the enhancement of anti-proliferative activities. In conclusion, the introduction of the 8-lipo group into aconitine leads to significant increase of anti-proliferative activity against MCF-7 and MCF-7/ADR cells, which suggests these kinds of lipo-alkaloids are powerful and promising antitumor compounds for breast cancer, especially for drug-resistant breast cancer.
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Antineoplásicos/farmacología , ADN-Topoisomerasas de Tipo II/metabolismo , Alcaloides Diterpénicos/química , Alcaloides Diterpénicos/farmacología , Inhibidores de Topoisomerasa II/farmacología , Antineoplásicos/química , Neoplasias de la Mama , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , ADN-Topoisomerasas de Tipo II/genética , Doxorrubicina/farmacología , Diseño de Fármacos , Etopósido/farmacología , Femenino , Humanos , Estructura Molecular , Relación Estructura-ActividadRESUMEN
The stereoselective fates of chiral pesticides in the environment has been reported in many studies. However, there is little data focused on the fate of chiral fosthiazate in the soil and aquatic ecosystems at chiral view. This study investigated the stereoselective fate of fosthiazate in the soil and aquatic ecosystems using ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) and liquid chromatography tandem time-of-flight mass spectrometry (LC-TOF/MS/MS). Significant stereoselective degradation among four fosthiazate stereoisomers were found in both greenhouse soil and water-sediment microcosms. In greenhouse soil, (1R,3S)-fosthiazate degraded faster than other three stereoisomers with the half-life of 2.7 d. The fosthiazate stereisomers in the seawater-sediment microcosm degraded more rapidly than in the river water-sediment microcosm. However, (1S,3R)-fosthiazate and (1S,3S)-fosthiazate possessed shorter degradation half-lives than their enantiomers in both microcosms, with the half-lives ranging from 3.4 d to 15.8 d. Ten degradation products were identified in the water-sediment microcosms, and, six of them were reported for the first time. Oxidation and hydrolysis were confirmed as the main degradation pathways of fosthiazate in the water-sediment microcosms. Our results revealed that the (1R,3S)-fosthiazate and (1R,3R)-fosthiazate may cause more serious ecotoxicity due to the longer half-lives than the other two stereoisomers in environment.
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Suelo , Agua , Cromatografía Liquida , Ecosistema , Compuestos Organofosforados , Espectrometría de Masas en Tándem , TiazolidinasRESUMEN
Pydiflumetofen is a novel and efficient broad-spectrum chiral fungicide consisting of a pair of enantiomers. A simple and sensitive chiral analytical method was established to determine the enantiomers of this chiral fungicide in food and environmental samples by ultra-high-performance liquid chromatography tandem triple quadrupole mass spectrometry (UHPLC-MS/MS) using QuEChERS method coupled with octadecylsilane-dispersive solid-phase extraction (C18-dSPE) as extraction procedure. The specific optical rotation and the absolute configuration of the enantiomers were identified by polarimetry and electronic circular dichroism (ECD). The elution order of the pydiflumetofen enantiomers on Lux Cellulose-2 was S-(-)-pydiflumetofen and R-(+)-pydiflumetofen. The average recoveries of eleven matrices ranged from 71.3% to 107.4%. The intraday relative standard deviations (RSDs) were less than 11.8%, and the interday RSDs were less than 12.6% for the two enantiomers. Stereoselective dissipation in pakchoi and soil were observed: S-(-)-pydiflumetofen was degraded faster than R-(+)-pydiflumetofen in pakchoi, causing the enantiomer fraction (EF) of the enantiomers to change from 0.50 to 0.42 in 7 days. However, R-(+)-pydiflumetofen was degraded faster than S-(-)-pydiflumetofen in soil, causing the EF of the enantiomers to change from 0.49 to 0.52 in 21 days. This study provides a method for monitoring pydiflumetofen enantiomer residues, which is crucial for improving risk assessments and the development of chiral pesticides.
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Fungicidas Industriales/análisis , Pirazoles/análisis , Cromatografía Líquida de Alta Presión/métodos , Cromatografía Liquida , Plaguicidas/análisis , Suelo/química , Contaminantes del Suelo/análisis , Extracción en Fase Sólida/métodos , Estereoisomerismo , Espectrometría de Masas en Tándem/métodosRESUMEN
BACKGROUND: The basic helix-loop-helix (bHLH) is the second largest gene family in the plant, some members play important roles in pistil development and response to drought, waterlogging, cold stress and salt stress. The bHLH gene family has been identified in many species, except for Brassica oleracea and B. napus thus far. This study aims to identify the bHLH family members in B. oleracea, B. rapa and B. napus, and elucidate the expression, duplication, phylogeny and evolution characters of them. RESULT: A total of 268 bHLH genes in B. oleracea, 440 genes in B. napus, and 251 genes in B. rapa, including 21 new bHLH members, have been identified. Subsequently, the analyses of the phylogenetic trees, conserved motifs and gene structures showed that the members in the same subfamily were highly conserved. Most Ka/Ks values of homologous gene were < 1, which indicated that these genes suffered from strong purifying selection for retention. The retention rates of BrabHLH and BolbHLH genes were 51.6 and 55.1%, respectively. The comparative expression patterns between B. rapa and B. napus showed that they had similar expression patterns in the root and contrasting patterns in the stems, leaves, and reproductive tissues. In addition, there were 41 and 30 differential expression bHLH genes under the treatments of ABA and JA, respectively, and the number of down regulation genes was significantly more than up regulation genes. CONCLUSION: In the present study, we identified and performed the comparative genomics analysis of bHLH gene family among B. oleracea, B. rapa and B. napus, and also investigated their diversity. The expression patterns between B. rapa and B. napus shows that they have the similar expression pattern in the root and opposite patterns in the stems, leaves, and reproduction tissues. Further analysis demonstrated that some bHLH gene members may play crucial roles under the abiotic and biotic stress conditions. This is the first to report on the bHLH gene family analysis in B. oleracea and B. napus, which can offer useful information on the functional analysis of the bHLH gene in plants.
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Brassica/genética , Evolución Molecular , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Secuencias Hélice-Asa-Hélice , Brassica napus/genética , Brassica rapa/genética , Duplicación de Gen , Perfilación de la Expresión Génica , Genoma de Planta , Genómica , FilogeniaRESUMEN
Real-time in situ imaging of organelles is increasingly important in modern biomedical analysis and diseases diagnosis. To realize this goal, organelle-targeting nanoparticles as one of the most commonly used technologies in subcellular sensing and imaging has attracted a lot of interest. The biocompatibility, specificity, and binding efficiency are especially critical for efficient organelle-targeting bioimaging. Gold nanoparticles (AuNPs) fabricated with bifunctional peptides constructed with both Au-binding affinity and nucleus-targeting ability were designed and examined for efficient nucleus-targeting bioimaging. Such a design is expected to achieve an oriented assembling of peptides by the medium of the Au-binding peptides specifically assembled on the surface of AuNPs, with the nucleus-targeting end open for accessibility. The bifunctional peptides showed strong binding affinity toward AuNPs and led to a binding capability â¼1.5 times higher than that of the bare nucleus-targeting peptides, ensuring good surface coverage of the nanoparticles for enhanced nucleus-targeting ability. Such fabricated AuNPs demonstrated over 90% cell viability after incubation for 24 h with HepG2 cells, which were highly biocompatible. Precise and efficient bioimaging of the nucleus was achieved for HepG2 cells by using the fabricated AuNPs as observed with a confocal laser scanning microscope, a dark-field/fluorescence microscope, and a transmission electron microscope. The high surface coverage and oriented binding pattern appeared to be a promising strategy for construction of organelle-targeting agencies.
Asunto(s)
Núcleo Celular/química , Oro/química , Nanopartículas del Metal/química , Péptidos/química , Núcleo Celular/metabolismo , Supervivencia Celular/efectos de los fármacos , Oro/farmacología , Humanos , Tamaño de la Partícula , Péptidos/farmacología , Espectrometría de Fluorescencia , Propiedades de Superficie , Células Tumorales CultivadasRESUMEN
BACKGOUND: Although Myeloid-derived suppressor cells (MDSCs) have a prominent ability to suppress the immune responses of T lymphocytes and propel tumor immune escape, a lack of profound systemic immunesuppression in tumor-bearing mice and tumor patients. The underlying mechanism of these remains unclear. METHODS: For this purpose, renal cancer-derived exosomes (RDEs) were first labeled with PKH67 and been observed the internalization by MDSCs. Flow cytometry analysis showed the proportion and activity change of MDSCs in spleen and bone marrow induced by RDEs. Further, western blot experiments were used to verify triggered mechanism of MDSCs by RDEs. Finally, proliferation and cytotoxicity of cytotoxic T lymphocytes (CTLs) co-cultured with MDSCs in vitro and a series of experiments in vivo were performed to demonstrate the specific inhibitory effect of RDEs-induced MDSCs. RESULTS: This study suggested that RDEs crucially contributed to presenting antigenic information, activating and driving specific immunosuppressive effect to MDSCs. HSP70, which is highly expressed in RDEs, initiate this process in a toll like receptor 2 (TLR2)-dependent manner. Importantly, RDEs-induced MDSCs could exert an antigen-specific immunosuppression effect on CTL and specific promote renal tumors-growth and immune escape in consequence. CONCLUSION: The immunosuppression mediated by MDSCs which is induced by RDEs is antigen-specific. HSP70, which is highly expressed in RDEs, plays a pivotal role in this process. Targeted abrogating the function of MDSCs, or eliminating the expression of HSP70 in exosomes, or blocking the crosstalk between them provides a new direction and theoretical support for future immunotherapy. Video abstract.