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Helicostoa sinensis E. Lamy, 1926 is a unique freshwater gastropod species with a sessile habit. This enigmatic species was first found cemented on river limestones from China about 120 years ago and described together with the genus. It was never collected again and has been considered monotypic. Here, we report the rediscovery of Helicostoa from several rivers in China, and describe a second species of this genus based on a comprehensive study. In addition to the unique sessile habit of both species, the new Helicostoa species presents one of the most remarkable cases of sexual dimorphism within molluscs. Only the adult female is sessile and the original aperture of the female is sealed by shell matter or rock, while an opening on the body whorl takes the function of the original aperture. The male is vagile, with a normal aperture. Our results confirm the recently suggested placement of Helicostoa within the family Bithyniidae. The sessility of Helicostoa species is considered as an adaption to the limestone habitat in large rivers. The extreme sexual dimorphism and secondary aperture of females are considered as adaptations to overcome the obstacles for mating and feeding that come with a sessile life style.
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Água Doce , Caracteres Sexuais , Feminino , Masculino , Animais , Rios , Carbonato de Cálcio , CaramujosRESUMO
The hitherto unknown hexakis(halomethyl)-functionalized tribenzotriquinacenes (TBTQs) 9 and 10 were synthesized from the key 4b,8b,12b-tribromo-TBTQ derivative 6 by an improved route in 67% overall yield. Extension of the bowl-shaped framework of 9 or 10 by threefold condensation with propargylamine or 2-azidoethylamine afforded the corresponding TBTQ-trialkyne 11 and TBTQ-triazide 12, respectively. While attempts to construct bis-TBTQ cages, including homodimerization of 11 and heterocoupling of 11 with 12, were unsuccessful, triazide 12 was found to undergo threefold [3 + 2]-cycloaddition with 3-ethynylaniline and phloroglucinol tripropargyl ether under click chemistry conditions. The latter reaction enabled facile capping of the TBTQ bowl to give the novel cage compound 5 in 22% yield.
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PANoptosis is a new type of cell death featured with pyroptosis, apoptosis and necroptosis, and is implicated in organ injury and mortality in various inflammatory diseases, such as sepsis and hemophagocytic lymphohistiocytosis (HLH). Reverse electron transport (RET)-mediated mitochondrial reactive oxygen species (mtROS) has been shown to contribute to pyroptosis and necroptosis. In this study we investigated the roles of mtROS and RET in PANoptosis induced by TGF-ß-activated kinase 1 (TAK1) inhibitor 5Z-7-oxozeaenol (Oxo) plus lipopolysaccharide (LPS) as well as the effects of anti-RET reagents on PANoptosis. We showed that pretreatment with anti-RET reagents 1-methoxy PMS (MPMS) or dimethyl fumarate (DMF) dose-dependently inhibited PANoptosis in macrophages BMDMs and J774A.1 cells induced by Oxo/LPS treatment assayed by propidium iodide (PI) staining. The three arms of the PANoptosis signaling pathway, namely pyroptosis, apoptosis and necroptosis signaling, as well as the formation of PANoptosomes were all inhibited by MPMS or DMF. We demonstrated that Oxo/LPS treatment induced RET and mtROS in BMDMs, which were reversed by MPMS or DMF pretreatment. Interestingly, the PANoptosome was co-located with mitochondria, in which the mitochondrial DNA was oxidized. MPMS and DMF fully blocked the mtROS production and the formation of PANoptosome induced by Oxo plus LPS treatment. An HLH mouse model was established by poly(I:C)/LPS challenge. Pretreatment with DMF (50 mg·kg-1·d-1, i.g. for 3 days) or MPMS (10 mg·kg-1·d-1, i.p. for 2 days) (DMF i.g. MPMS i.p.) effectively alleviated HLH lesions accompanied by decreased hallmarks of PANoptosis in the liver and kidney. Collectively, RET and mtDNA play crucial roles in PANoptosis induction and anti-RET reagents represent a novel class of PANoptosis inhibitors by blocking oxidation of mtDNA, highlighting their potential application in treating PANoptosis-related inflammatory diseases. PANoptotic stimulation induces reverse electron transport (RET) and reactive oxygen species (ROS) in mitochondia, while 1-methoxy PMS and dimethyl fumarate can inhibit PANoptosis by suppressing RETmediated oxidation of mitochondrial DNA.
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DNA Mitocondrial , Fumarato de Dimetilo , Animais , Camundongos , Espécies Reativas de Oxigênio/metabolismo , Transporte de Elétrons , Fumarato de Dimetilo/metabolismo , Fumarato de Dimetilo/farmacologia , DNA Mitocondrial/metabolismo , Lipopolissacarídeos/farmacologia , Elétrons , Mitocôndrias , ApoptoseRESUMO
Reported herein is a Paternò-Büchi reaction of aromatic double bonds with quinones under visible light irradiation. The reactions of aromatics with quinones exposed to blue LED irradiation yielded oxetanes at -78 °C, which was attributed to both the activation of double bonds in aromatics and the stabilization of oxetanes by thiadiazole, oxadiazole, or selenadiazole groups. The addition of Cu(OTf)2 to the reaction system at room temperature resulted in the formation of diaryl ethers via the copper-catalyzed ring opening of oxetanes in situ. Notably, the substrate scope was extended to general aromatics.
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Macrophages represent the first lines of innate defense against pathogenic infections and are poised to undergo multiple forms of regulated cell death (RCD) upon infections or toxic stimuli, leading to multiple organ injury. Triptolide, an active compound isolated from Tripterygium wilfordii Hook F., possesses various pharmacological activities including anti-tumor and anti-inflammatory effects, but its applications have been hampered by toxic adverse effects. It remains unknown whether and how triptolide induces different forms of RCD in macrophages. In this study, we showed that triptolide exhibited significant cytotoxicity on cultured macrophages in vitro, which was associated with multiple forms of lytic cell death that could not be fully suppressed by any one specific inhibitor for a single form of RCD. Consistently, triptolide induced the simultaneous activation of pyroptotic, apoptotic and necroptotic hallmarks, which was accompanied by the co-localization of ASC specks respectively with RIPK3 or caspase-8 as well as their interaction with each other, indicating the formation of PANoptosome and thus the induction of PANoptosis. Triptolide-induced PANoptosis was associated with mitochondrial dysfunction and ROS production. PANoptosis was also induced by triptolide in mouse peritoneal macrophages in vivo. Furthermore, triptolide caused kidney and liver injury, which was associated with systemic inflammatory responses and the activation of hallmarks for PANoptosis in vivo. Collectively, our data reveal that triptolide induces PANoptosis in macrophages in vitro and exhibits nephrotoxicity and hepatotoxicity associated with induction of PANoptosis in vivo, suggesting a new avenue to alleviate triptolide's toxicity by harnessing PANoptosis.
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Diterpenos , Fenantrenos , Camundongos , Animais , Apoptose , Macrófagos/metabolismo , Diterpenos/efeitos adversos , Diterpenos/metabolismo , Fenantrenos/toxicidade , Fenantrenos/metabolismo , Compostos de Epóxi/toxicidade , Compostos de Epóxi/metabolismoRESUMO
BACKGROUND: Haemophilus influenzae (H. influenzae), Streptococcus pneumoniae (pneumococcus) and influenza vaccines are administered in children to prevent infections caused by these pathogens. The benefits of vaccination for asthma control in children and the elicited immune response are not fully understood. This study aimed to investigate the impact of these vaccinations on respiratory infections, asthma symptoms, asthma severity and control status, pathogen colonization and in vitro immune responses to different stimulants mimicking infections in asthmatic children. METHODS: Children aged 4-6 years were recruited into the multicentre prospective PreDicta study conducted across five European countries. Information about vaccination history, infections, antibiotic use, inhaled corticosteroid (ICS) use and asthma symptoms in the last 12 months were obtained from questionnaires of the study. Nasopharyngeal samples were collected at the first visit to assess bacterial and viral colonization, and venous blood for isolation of peripheral blood mononuclear cells (PBMCs). The PBMCs were stimulated with phytohemagglutinin, R848, Poly I:C and zymosan. The levels of 22 cytokines and chemokines were measured in cell culture supernatants using a luminometric multiplex assay. RESULTS: One-hundred and forty asthmatic preschool children (5.3 ± 0.7 years) and 53 healthy children (5.0 ± 0.8 years) from the PreDicta cohort were included in the current study. Asthmatic children were associated with more frequent upper and lower respiratory infections, and more frequent and longer duration of antibiotic use compared with healthy children. In asthmatic children, sufficient H. influenzae vaccination was associated with a shorter duration of upper respiratory infection (URI) and overall use and average dose of ICS. The airway colonization was characterized by less pneumococcus and more rhinovirus. Pneumococcal vaccination was associated with a reduction in the use rate and average dose of ICS, improved asthma control, and less human enterovirus and more H. influenzae and rhinovirus (RV) airway colonization. Influenza vaccination in the last 12 months was associated with a longer duration of URI, but with a decrease in the occurrence of lower respiratory infection (LRI) and the duration of gastrointestinal (GI) infection and antibiotic use. Asthmatic preschoolers vaccinated with H. influenzae, pneumococcus or influenza presented higher levels of Th1-, Th2-, Th17- and regulatory T cells (Treg)-related cytokines in unstimulated PBMCs. Under stimulation, PBMCs from asthmatic preschoolers with pneumococcal vaccination displayed a predominant anti-inflammatory immune response, whereas PBMCs from asthmatic children with sufficient H. influenzae or influenza vaccination were associated with both pro- and anti-inflammatory immune responses. CONCLUSION: In asthmatic preschoolers, the standard childhood vaccinations to common respiratory pathogens have beneficial effects on asthma control and may modulate immune responses relevant to asthma pathogenesis.
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Asma , Influenza Humana , Infecções Respiratórias , Humanos , Pré-Escolar , Lactente , Streptococcus pneumoniae , Haemophilus influenzae , Influenza Humana/prevenção & controle , Estudos Prospectivos , Leucócitos Mononucleares , Infecções Respiratórias/microbiologia , Citocinas , Imunidade , Vacinação , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Anti-InflamatóriosRESUMO
Necroptosis has been implicated in various inflammatory diseases including tumor-necrosis factor-α (TNF-α)-induced systemic inflammatory response syndrome (SIRS). Dimethyl fumarate (DMF), a first-line drug for treating relapsing-remitting multiple sclerosis (RRMS), has been shown to be effective against various inflammatory diseases. However, it is still unclear whether DMF can inhibit necroptosis and confer protection against SIRS. In this study, we found that DMF significantly inhibited necroptotic cell death in macrophages induced by different necroptotic stimulations. Both the autophosphorylation of receptor-interacting serine/threonine kinase 1 (RIPK1) and RIPK3 and the downstream phosphorylation and oligomerization of MLKL were robustly suppressed by DMF. Accompanying the suppression of necroptotic signaling, DMF blocked the mitochondrial reverse electron transport (RET) induced by necroptotic stimulation, which was associated with its electrophilic property. Several well-known anti-RET reagents also markedly inhibited the activation of the RIPK1-RIPK3-MLKL axis accompanied by decreased necrotic cell death, indicating a critical role of RET in necroptotic signaling. DMF and other anti-RET reagents suppressed the ubiquitination of RIPK1 and RIPK3, and they attenuated the formation of necrosome. Moreover, oral administration of DMF significantly alleviated the severity of TNF-α-induced SIRS in mice. Consistent with this, DMF mitigated TNF-α-induced cecal, uterine, and lung damage accompanied by diminished RIPK3-MLKL signaling. Collectively, DMF represents a new necroptosis inhibitor that suppresses the RIPK1-RIPK3-MLKL axis through blocking mitochondrial RET. Our study highlights DMF's potential therapeutic applications for treating SIRS-associated diseases.
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Proteínas Quinases , Fator de Necrose Tumoral alfa , Camundongos , Animais , Fator de Necrose Tumoral alfa/metabolismo , Proteínas Quinases/metabolismo , Fumarato de Dimetilo , Necroptose , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Síndrome de Resposta Inflamatória Sistêmica , Fosforilação Oxidativa , ApoptoseRESUMO
Activation of NLR family pyrin domain-containing 3 (NLRP3) inflammasome plays important role in defending against infections, but its aberrant activation is causally linked to many inflammatory diseases, thus being a therapeutic target for these diseases. Theaflavin, one major ingredient of black tea, exhibits potent anti-inflammatory and anti-oxidative activities. In this study, we investigated the therapeutic effects of theaflavin against NLRP3 inflammasome activation in macrophages in vitro and in animal models of related diseases. We showed that theaflavin (50, 100, 200 µM) dose-dependently inhibited NLRP3 inflammasome activation in LPS-primed macrophages stimulated with ATP, nigericin or monosodium urate crystals (MSU), evidenced by reduced release of caspase-1p10 and mature interleukin-1ß (IL-1ß). Theaflavin treatment also inhibited pyroptosis as shown by decreased generation of N-terminal fragment of gasdermin D (GSDMD-NT) and propidium iodide incorporation. Consistent with these, theaflavin treatment suppressed ASC speck formation and oligomerization in macrophages stimulated with ATP or nigericin, suggesting reduced inflammasome assembly. We revealed that theaflavin-induced inhibition on NLRP3 inflammasome assembly and pyroptosis resulted from ameliorated mitochondrial dysfunction and reduced mitochondrial ROS production, thereby suppressing interaction between NLRP3 and NEK7 downstream of ROS. Moreover, we showed that oral administration of theaflavin significantly attenuated MSU-induced mouse peritonitis and improved the survival of mice with bacterial sepsis. Consistently, theaflavin administration significantly reduced serum levels of inflammatory cytokines including IL-1ß and attenuated liver inflammation and renal injury of mice with sepsis, concomitant with reduced generation of caspase-1p10 and GSDMD-NT in the liver and kidney. Together, we demonstrate that theaflavin suppresses NLRP3 inflammasome activation and pyroptosis by protecting mitochondrial function, thus mitigating acute gouty peritonitis and bacterial sepsis in mice, highlighting a potential application in treating NLRP3 inflammasome-related diseases.
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Gota , Peritonite , Sepse , Camundongos , Animais , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Espécies Reativas de Oxigênio , Nigericina/uso terapêutico , Peritonite/tratamento farmacológico , Antioxidantes/uso terapêutico , Sepse/complicações , Sepse/tratamento farmacológico , Caspases , Trifosfato de Adenosina , Interleucina-1beta/metabolismoRESUMO
BACKGROUND: As an anticancer Chinese herbal medicine, the effective components and mechanism of Actinidia chinensis Planch (ACP, Tengligen) in the treatment of colon cancer are still unclear. In the present study, the integration of network pharmacology, molecular docking, and cell experiments was employed to study the effective mechanism of ACP against colon cancer. METHODS: The Venn diagram and STRING database were used to construct the protein-protein interaction network (PPI) of ACP-colon cancer, and further topological analysis was used to obtain the key target genes of ACP in colon cancer. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were used to visualize the related functions and pathways. Molecular docking between key targets and compounds was determined using software such as AutoDockTools. Finally, the effect of ACP on CT26 cells was observed in vitro. RESULTS: The study identified 40 ACP-colon key targets, including CASP3, CDK2, GSK3B, and PIK3R1. GO and KEGG enrichment analyses found that these genes were involved in 211 biological processes and 92 pathways, among which pathways in cancer, PI3K-Akt, p53, and cell cycle might be the main pathways of ACP against colon cancer. Molecular docking verified that the key components of ACP could stably bind to the corresponding targets. The experimental results showed that ACP could inhibit proliferation, induce apoptosis, and downregulate the phosphorylation of PIK3R1, Akt, and GSK3B in CT26 cells. CONCLUSION: ACP is an anti-colon cancer herb with multiple components, and involvement of multiple target genes and signaling pathways. ACP can significantly inhibit proliferation and induce apoptosis of colon cancer cells, which may be closely related to the regulation of PI3K/AKT/GSK3B signal transduction.
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Actinidia , Neoplasias do Colo , Simulação de Acoplamento Molecular , Actinidia/genética , Farmacologia em Rede , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/genética , Fatores de TranscriçãoRESUMO
Moderate stimuli in mitochondria improve wide-ranging stress adaptability in animals, but whether mitochondria play similar roles in plants is largely unknown. Here, we report the enhanced stress adaptability of S-type cytoplasmic male sterility (CMS-S) maize and its association with mild expression of sterilizing gene ORF355. A CMS-S maize line exhibited superior growth potential and higher yield than those of the near-isogenic N-type line in saline fields. Moderate expression of ORF355 induced the accumulation of reactive oxygen species and activated the cellular antioxidative defense system. This adaptive response was mediated by elevation of the nicotinamide adenine dinucleotide concentration and associated metabolic homeostasis. Metabolome analysis revealed broad metabolic changes in CMS-S lines, even in the absence of salinity stress. Metabolic products associated with amino acid metabolism and galactose metabolism were substantially changed, which underpinned the alteration of the antioxidative defense system in CMS-S plants. The results reveal the ORF355-mediated superior stress adaptability in CMS-S maize and might provide an important route to developing salt-tolerant maize varieties.
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Infertilidade das Plantas , Zea mays , Zea mays/genética , Infertilidade das Plantas/genética , Mitocôndrias/metabolismo , Citoplasma/metabolismo , HomeostaseRESUMO
This study was aimed to investigate the effect of hypoxia on lipopolysaccharide (LPS)-induced CXC-chemokine ligand-10 (CXCL10) expression and the underlying mechanism. C57BL/6J mice were randomly divided into control, hypoxia, LPS, and hypoxia combined with LPS groups. The LPS group was intraperitoneally injected with 0.5 mg/kg LPS, and the hypoxia group was placed in a hypobaric hypoxia chamber (simulated altitude of 6 000 m). The serum and hippocampal tissue samples were collected after 6 h of the treatment. The levels of CXCL10 in the serum and hippocampal tissue of mice were detected by ELISA. The microglia cell line BV2 and primary microglia were stimulated with hypoxia (1% O2) and/or LPS (100 ng/mL) for 6 h. The mRNA expression level of CXCL10 and its content in culture supernatant were detected by real-time quantitative PCR and ELISA, respectively. The phosphorylation levels of nuclear factor κB (NF-κB) signaling pathway-related proteins, p65 and IκBα, were detected by Western blot. Moreover, after NF-κB signaling pathway being blocked with a small molecular compound, PDTC, CXCL10 mRNA expression level was detected in the BV2 cells. The results showed that in the LPS-induced mouse inflammatory model, hypoxia treatment could promote LPS-induced up-regulation of CXCL10 in both serum and hippocampus. Compared with the cells treated with LPS alone, the expression of CXCL10 mRNA and the content of CXCL10 in the culture supernatant of BV2 cells treated with hypoxia combined with LPS were significantly increased. The CXCL10 mRNA level of primary microglial cells treated with hypoxia combined with LPS was significantly up-regulated. Compared with the cells treated with hypoxia or LPS alone, the phosphorylation levels of p65 and IκBα in the BV2 cells treated with hypoxia combined with LPS were significantly increased. PDTC blocked the induction of CXCL10 gene expression by LPS in the BV2 cells. These results suggest that hypoxia promotes LPS-induced expression of CXCL10 in both animal and cell models, and NF-κB signaling pathway plays an important role in this process.
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Microglia , NF-kappa B , Animais , Camundongos , Quimiocinas CXC/metabolismo , Quimiocinas CXC/farmacologia , Hipóxia , Ligantes , Lipopolissacarídeos/farmacologia , Camundongos Endogâmicos C57BL , Microglia/metabolismo , NF-kappa B/metabolismo , Inibidor de NF-kappaB alfa/metabolismo , Inibidor de NF-kappaB alfa/farmacologia , RNA Mensageiro/metabolismoRESUMO
As a metal-free photocatalyst, graphitic carbon nitride (g-CN) shows great potential for photocatalytic water splitting, although its performance is significantly limited by structural defects due to incomplete polymerization. In the present work, we successfully synthesize highly conjugated g-CN nanofoam through an iodide substitution technique. The product possesses a high polymerization degree, low defect density, and large specific surface area; as a result, it achieves a hydrogen evolution rate of 9.06 mmol h-1 g-1 under visible light irradiation, with an apparent quantum efficiency (AQE) of 18.9% at 420 nm. Experimental analysis and theoretical calculations demonstrate that the recombination of photogenerated carriers at C-NHx defects was effectively depressed in the nanofoam, giving rise to the high photocatalytic activity.
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BACKGROUND: Many patients with neurological disorders experience chronic fatigue, but the neural mechanisms involved are unclear. OBJECTIVE: Here we investigated whether the brain structural and functional connectivity alterations were involved in fatigue related to neuromyelitis optica spectrum disorder (NMOSD). METHODS: This prospective pilot study used structural and resting-state functional brain magnetic resonance imaging to compare total cortical thickness, cortical surface area, deep gray matter volume and functional connectivity (FC) between 33 patients with NMOSD and 20 healthy controls (HCs). Patients were subgrouped as low fatigue (LF) and high fatigue (HF). RESULTS: HF patients scored higher on the Hamilton Anxiety Rating Scale and Hamilton Rating Scale for Depression than LF patients and HCs. The two patient subgroups and HC group did not differ significantly in cortical thickness, cortical surface area and volumes of the bilateral caudate nucleus, bilateral putamen, bilateral amygdala, bilateral hippocampus, bilateral thalamus proper or right nucleus accumbens (p > 0.05). However, after correcting for age, sex, years of education, anxiety and depression, HF patients showed larger left pallidum than HCs (0.1573 ± 0.0214 vs 0.1372 ± 0.0145, p = 0.009). Meanwhile, both LF patients (0.0377 ± 0.0052 vs 0.0417 ± 0.0052, p = 0.009) and HF patients (0.0361 ± 0.0071 vs 0.0417 ± 0.0052, p = 0.013) showed smaller left nucleus accumbens than HCs.. Compared with LF patients, HF patients showed significantly decreased FC between the left pallidum and bilateral cerebellar posterior lobes. CONCLUSIONS: This was the first evidence linking structural and functional alterations in the brain to fatigue in NMOSD, and in the future, long term follow-up was necessary.
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Neuromielite Óptica , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Humanos , Imageamento por Ressonância Magnética/métodos , Neuromielite Óptica/complicações , Neuromielite Óptica/diagnóstico por imagem , Neuromielite Óptica/patologia , Projetos Piloto , Estudos ProspectivosRESUMO
Male sterility refers to the defective development of male reproductive organs, which led to plants incapable of producing normal and functional pollens. Maize (Zea mays L.) is one of the most important food crops, as well as one of the earliest crops to utilize heterosis in breeding. Single cross hybrid has been the main type of maize heterosis utilization for a long time. The planting area of maize hybrid in China has been stable at about 620 million mu. More than one billion kilograms of commercial hybrid seeds are needed each year, and the annual seed production area has been stable at about 2.5 million mu in recent years. So far, manual emasculation has been the major way of maize hybrid seed production in China, which is laborious and time consuming. Generally, spatial isolation is necessary for maize hybrid seed production, this requirement results in only some regions in the country suitable for maize hybrid seed production. Manual emasculation requires seasonal demand of labors. At present, with the urbanization of a large number of rural laborers, the seed production regions experience a serious labor shortage. Accordingly, the cost of seed production increases with the rising of land rent and labor costs. In addition, it is difficult to guarantee the seed purity with manual or mechanical emasculation for hybrid seed production. However, incorporating male sterility into maize hybrid seed production could reduce its cost and ensure hybrid seed purity. It can also avoid the difficulties of manual or mechanical emasculation in field operation under extreme weather conditions. Therefore, it is the inevitable trend of development in the maize seed industry. In this review, we summarize the exploitation and creation of maize cytoplasmic male sterility (CMS), maize genic male sterility (GMS) resources in China, and the developing process from natural discovery to targeted creation of male sterility resources in plants, and the research progress of maize male sterility. We then analyze the application status and existing problems of maize male sterility, based on the development trend of maize seed industry, as well as the research and application status of male sterility in China. We also identify seven aspects that need to be further strengthen, thereby providing the reference for the creation, research and utilization of maize male sterility in the future.
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Infertilidade Masculina , Zea mays , Produtos Agrícolas/genética , Melhoramento Vegetal , Infertilidade das Plantas/genética , Sementes/genética , Zea mays/genéticaRESUMO
Nitric oxide (NO) is a molecule of physiological importance, and the function of NO depends on its concentration in biological systems, particularly in cells. Concentration-based analysis of intracellular NO can provide insight into its precise role in health and disease. However, current methods for detecting intracellular NO are still inadequate for quantitative analysis. In this study, we report a quantitative mass spectrometry probe approach to measure NO levels in cells. The probe, Amlodipine (AML), comprises a Hantzsch ester group that reacts with NO to form a pyridine, Dehydro Amlodipine (DAM). Quantification of DAM by ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) allows specific measurement of intracellular NO levels. Notably, the AML/NO reaction proceeds rapidly (within 1 s), which is favorable for NO detection considering its large diffusivity and short half-life. Meanwhile, studies under simulated physiological conditions revealed that the AML response to NO is proportional and selective. The presented UPLC-MS/MS method showed high sensitivity (LLOQ = 0.24 nM) and low matrix interference (less than 15%) in DAM quantification. Furthermore, the mass spectrometry probe approach was demonstrated by enabling the measurement of endogenous and exogenous NO in cells. Hence, the quantitative UPLC-MS/MS method developed using AML as a probe is expected to be a new method for intracellular NO analysis.
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Óxido Nítrico , Espectrometria de Massas em Tandem , Cromatografia Líquida de Alta Pressão , Cromatografia Líquida , Reprodutibilidade dos TestesRESUMO
Getah virus (GETV), a mosquito-borne virus belonging to the Alphavirus genus of family Togaviridae, has become increasingly problematic, which poses a huge threat to the safety of animals and public health. In order to detect GETV quickly and accurately, we have developed a SYBR Green I real-time quantitative reverse transcription PCR (RT-qPCR) assay for GETV with the detection limit of 66 copies/µL, excellent correlation coefficient (R2) of 0.9975, and amplification efficiency (E) of 98.90%, the target selected was the non-structural protein 3 of GETV. The sensitivity of it was higher than that of ordinary RT-PCR by 1000 folds, and the inter-assay and intra-assay CV values were all less than 0.99%. The newly developed RT-qPCR assay exhibited good sensitivity and reproducibility, which will provide technical support for the reliable and specific rapid diagnosis, and quantitative analysis of GETV infection.
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Alphavirus , Culicidae , Alphavirus/genética , Animais , Benzotiazóis , Diaminas , Quinolinas , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Transcrição Reversa , Sensibilidade e Especificidade , SuínosRESUMO
BACKGROUND: Stalk fracture caused by strong wind can severely reduce yields in maize. Stalks with higher stiffness and flexibility will exhibit stronger lodging resistance. However, stalk flexibility is rarely studied in maize. Stalk fracture of the internode above the ear before tasseling will result in the lack of tassel and pollen, which is devastating for pollination in seed production. In this study, we focused on stalk lodging before tasseling in two maize inbred lines, JING724 and its improved line JING724A1 and their F2:3 population. RESULTS: JING724A1 showed a larger stalk fracture angle than JING724, indicating higher flexibility. In addition, compared to JING724, JING724A1 also had longer and thicker stalks, with a conical, frustum-shaped internode above the ear. Microscopy and X-ray microcomputed tomography of the internal stalk architecture revealed that JING724A1 had more vascular bundles and thicker sclerenchyma tissue. Furthermore, total soluble sugar content of JING724A1, especially the glucose component, was substantially higher than in JING724. Using an F2:3 population derived from a JING724 and JING724A1 cross, we performed bulk segregant analysis for stalk fracture angle and detected one QTL located on Chr3: 14.00-19.28 Mb. Through transcriptome data analysis and ∆ (SNP-index), we identified two candidate genes significantly associated with high stalk fracture angle, which encode a RING/U-box superfamily protein (Zm00001d039769) and a MADS-box transcription factor 54 (Zm00001d039913), respectively. Two KASP markers designed from these two candidate genes also showed significant correlations with stalk fracture angle. CONCLUSIONS: The internode shape and glucose content are possibly correlated with stalk flexibility in maize. Two genes in the detected QTL are potentially associated with stalk fracture angle. These novel phenotypes and associated loci will provide a theoretical foundation for understanding the genetic mechanisms of lodging, and facilitate the selection of maize varieties with improved flexibility and robust lodging resistance.
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Parede Celular/química , Caules de Planta/anatomia & histologia , Caules de Planta/crescimento & desenvolvimento , Caules de Planta/genética , Zea mays/anatomia & histologia , Zea mays/crescimento & desenvolvimento , Zea mays/genética , Produtos Agrícolas/anatomia & histologia , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Cruzamentos Genéticos , Genes de Plantas , Variação Genética , Genótipo , Fenótipo , Locos de Características QuantitativasRESUMO
Arabidopsis thaliana ENO2 (AtENO2) plays an important role in plant growth and development. It encodes two proteins, a full-length AtENO2 and a truncated version, AtMBP-1, alternatively translated from the second start codon of the mRNA. The AtENO2 mutant (eno2- ) exhibited reduced leaf size, shortened siliques, a dwarf phenotype and higher sensitivity to abiotic stress. The objectives of this study were to analyze the regulatory network of the ENO2 gene in plant growth development and understand the function of AtENO2/AtMBP-1 to abiotic stresses. An eno2- /35S:AtENO2-GFP line and an eno2- /35S:AtMBP-1-GFP line of Arabidopsis were obtained. Results of sequencing by 454 GS FLX identified 578 upregulated and 720 downregulated differential expressed genes (DEGs) in a pairwise comparison (WT-VS-eno2- ). All the high-quality reads were annotated using the Gene Ontology (GO) terms. The DEGs with KEGG pathway annotations occurred in 110 pathways. The metabolic pathways and biosynthesis of secondary metabolites contained more DEGs. Moreover, the eno2- /35S:AtENO2-GFP line returned to the wild-type (WT) phenotype and was tolerant to drought and salt stresses. However, the eno2- /35S:AtMBP-1-GFP line was not able to recover the WT phenotype but it has a higher tolerance to drought and salt stresses. Results from this study demonstrate that AtENO2 is critical for the growth and development, and the AtMBP-1 coded by AtENO2 is important in tolerance of Arabidopsis to abiotic stresses.
Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/fisiologia , Secas , Estresse Salino , Proteínas de Transporte , Regulação da Expressão Gênica de Plantas , Plantas Geneticamente ModificadasRESUMO
As a natural plant source of artemisinin,a first-line drug against malaria,Artemisia annua directly affects the extraction process of artemisinin and the source of artemisinin. At present,traditional breeding methods combined with tissue culture are often used to breed high-yield artemisinin-containing new varieties of A. annua. However,the breeding method has the disadvantages of low efficiency and continuous selection. In this study,heavy ion beam irradiation technology was used to observe the specific germplasm resources of A. annua,and the morphological characteristics,agronomic traits and artemisinin content were used as indicators to observe the selection materials and materials. The cultivated new varieties were compared with trials and regional trials. In addition,the new variety of A. annua was identified by SRAP molecular marker technology. The results showed that the new variety of A. annua, " Kehao No.1",had an average yield of 235. 0 kg of dry leaf per mu,which was more than 20% higher than that of the control. Especially,the average artemisinin content was 2. 0%,which was 45% higher than that of the control,and the " Kehao No.1" has high anti-white powder disease,high-yield and high-quality new varieties. Therefore,mutagenic breeding of heavy ion beam irradiation can significantly improve the yield and artemisinin content of the " Kehao No. 1" and it has a good promotion value.
Assuntos
Artemisia annua/genética , Artemisininas/análise , Melhoramento Vegetal , Plantas Medicinais/genética , Artemisia annua/química , Íons Pesados , Mutagênese , Fenótipo , Plantas Medicinais/químicaRESUMO
Salt stress is a major abiotic factor limiting maize yield. To characterize the mechanism underlying maize salt tolerance, we compared the seedling root proteomes of salt-tolerant Jing724 and salt-sensitive D9H. The germination rate and growth parameter values (weight and length) were higher for Jing724 than for D9H under saline conditions. Using an iTRAQ-based method, we identified 513 differentially regulated proteins (DRPs), with 83 and 386 DRPs specific to Jing724 and D9H, respectively. In salt-stressed Jing724, the DRPs were primarily associated with the pentose phosphate pathway, glutathione metabolism, and nitrogen metabolism. Key DRPs, such as glucose-6-phosphate 1-dehydrogenase, NADPH-producing dehydrogenase, glutamate synthase, and glutamine synthetase, were identified based on pathway enrichment and protein-protein interaction analyses. Moreover, salt-responsive proteins in Jing724 seedlings were implicated in energy management, maintenance of redox homeostasis, detoxification of ammonia, regulation of osmotic homeostasis, stress defense and adaptation, biotic cross-tolerance, and regulation of gene expression. Quantitative analyses of superoxide dismutase activity, malondialdehyde content, relative electrolyte leakage, and proline content were consistent with the predicted changes based on DRP functions. Furthermore, changes in the abundance of eight representative DRPs were correlated with the corresponding mRNA levels. Our results may be useful for elucidating the molecular networks mediating salt tolerance.