RESUMO
CRISPR-Cas9 is widely used for genome editing, but its PAM sequence requirements limit its efficiency. In this study, we explore Faecalibaculum rodentium Cas9 (FrCas9) for plant genome editing, especially in rice. FrCas9 recognizes a concise 5'-NNTA-3' PAM, targeting more abundant palindromic TA sites in plant genomes than the 5'-NGG-3' PAM sites of the most popular SpCas9. FrCas9 shows cleavage activities at all tested 5'-NNTA-3' PAM sites with editing outcomes sharing the same characteristics of a typical CRISPR-Cas9 system. FrCas9 induces high-efficiency targeted mutagenesis in stable rice lines, readily generating biallelic mutants with expected phenotypes. We augment FrCas9's ability to generate larger deletions through fusion with the exonuclease, TREX2. TREX2-FrCas9 generates much larger deletions than FrCas9 without compromise in editing efficiency. We demonstrate TREX2-FrCas9 as an efficient tool for genetic knockout of a microRNA gene. Furthermore, FrCas9-derived cytosine base editors (CBEs) and adenine base editors (ABE) are developed to produce targeted C-to-T and A-to-G base edits in rice plants. Whole-genome sequencing-based off-target analysis suggests that FrCas9 is a highly specific nuclease. Expression of TREX2-FrCas9 in plants, however, causes detectable guide RNA-independent off-target mutations, mostly as single nucleotide variants (SNVs). Together, we have established an efficient CRISPR-FrCas9 system for targeted mutagenesis, large deletions, C-to-T base editing, and A-to-G base editing in plants. The simple palindromic TA motif in the PAM makes the CRISPR-FrCas9 system a promising tool for genome editing in plants with an expanded targeting scope.
Assuntos
Sistemas CRISPR-Cas , Edição de Genes , Genoma de Planta , Oryza , Edição de Genes/métodos , Genoma de Planta/genética , Oryza/genética , Proteína 9 Associada à CRISPR/metabolismo , Proteína 9 Associada à CRISPR/genéticaRESUMO
Melatonin (N-acetyl-5-methoxytryptamine) plays important roles in plant defences against a variety of biotic and abiotic stresses, including UV-B stress. Molecular mechanisms underlying functions of melatonin in plant UV-B responses are poorly understood. Here, we show that melatonin effect on molecular signalling pathways, physiological changes and UV-B stress resistance in Arabidopsis. Both exogenous and endogenous melatonin affected expression of UV-B signal transduction pathway genes. Experiments using UV-B signalling component mutants cop1-4 and hy5-215 revealed that melatonin not only acts as an antioxidant to promote UV-B stress resistance, but also regulates expression of several key components of UV-B signalling pathway, including ubiquitin-degrading enzyme (COP1), transcription factors (HY5, HYH) and RUP1/2. Our findings indicate that melatonin delays and subsequently enhances expression of COP1, HY5, HYH and RUP1/2, which act as central effectors in UV-B signalling pathway, thus regulating their effects on antioxidant systems to protect the plant from UV-B stress.
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Arabidopsis/efeitos da radiação , Melatonina/metabolismo , Transdução de Sinais , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/fisiologia , Peróxido de Hidrogênio/metabolismo , Malondialdeído/metabolismo , Plantas Geneticamente Modificadas , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/efeitos da radiação , Estresse Fisiológico , Raios Ultravioleta/efeitos adversosRESUMO
The superoxide dismutase 1 (SOD1) mutation is one of the most notable causes of amyotrophic lateral sclerosis (ALS), and modifying the mutant SOD1 gene is the best approach for the treatment of patients with ALS linked to the mutations in this gene. Clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas9)/sgRNA delivered by the adeno-associated virus (AAV) system is a powerful tool for genome editing in the central nervous system (CNS). Here, we tested the capacity of the AAV-SaCas9-sgRNA system to modify mutant SOD1 in SOD1G93A transgenic mice and found that AAV9-SaCas9-sgRNA5 deleted the SOD1 gene, improved the lifespan of SOD1G93A mice by 54.6%, and notably ameliorated the performance of ALS transgenic mice. An immunochemical analysis showed that the expression of mutant SOD1 was very weak in motor neurons expressing SaCas9-sgRNA5. Consequently, the area showing muscle atrophy was more notably restored in the group treated with SaCas9-sgRNA5 compared with the group treated with SaCas9-sgLacZ. In addition, deep sequencing did not show the indel mutation in the gene highly matched to sgRNA5. Hence, AAV9-SaCas9-sgRNA-based gene editing is a feasible potential treatment for patients with ALS linked to SOD1 mutations.
Assuntos
Esclerose Lateral Amiotrófica/terapia , Sistemas CRISPR-Cas , Deleção de Genes , Edição de Genes/métodos , Terapia Genética/métodos , Superóxido Dismutase-1/genética , Esclerose Lateral Amiotrófica/genética , Animais , Feminino , Células HEK293 , Humanos , Masculino , Camundongos , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Mutação de Sentido Incorreto , RNA Guia de Cinetoplastídeos/genética , RNA Guia de Cinetoplastídeos/metabolismo , Superóxido Dismutase-1/metabolismoRESUMO
Respiratory ailments have plagued occupational and public health communities exposed to World Trade Center (WTC) dust since the September 11, 2001 attack on the Twin Towers in Lower Manhattan. We proposed that these ailments were proposed to be induced by inhalation exposure to WTC particulate matter (WTCPM), that was released during the collapse of the buildings and its subsequent resuspension during cleanup. We investigated this hypothesis using both an in vitro and an in vivo mouse intranasal (IN) exposure models to identify the inflammatory potential of WTCPM with specific emphasis on respiratory and endothelial tissue responses. The in vitro exposure studies found WTCPM exposure to be positively correlated with cytotoxicity and increased NO2- production in both BEAS-2B pulmonary epithelial cells and THP-1 macrophage cells. The in vivo C57BL/6 mouse studies found significant increases in inflammatory markers including increases in polymorphonuclear neutrophil (PMN) influx into nasal and bronchoalveolar lavage fluids (NLF and BALF), as well as increased levels of total protein and cytokine/chemokines levels. Concurrently, NLF, BALF, and serum NO2- levels exhibited significant homeostatic temporal deviations as well as temporal myograohic aortic dysfunction in myography studies. Respiratory exposure to- and evidence -based retention of- WTCPM may have contributed to chronic systemic effects in exposed mice that r resembled to observed effects in WTCPM-exposed human populations. Collectively, these findings are reflective of WTCPM exposure and its effect(s) on respiratory and aortic tissues, highlighting potential dysfunctional pathways that may precipitate inflammatory events, while simultaneously altering homeostatic balances. The tight interplay between these balances, when chronically altered, may contribute to- or result in- chronically diseased pathological states.
Assuntos
Poluentes Atmosféricos/toxicidade , Materiais de Construção/toxicidade , Poeira/análise , Endotélio Vascular/efeitos dos fármacos , Exposição por Inalação/efeitos adversos , Pneumonia/induzido quimicamente , Poluentes Atmosféricos/análise , Animais , Aorta/efeitos dos fármacos , Aorta/fisiopatologia , Biomarcadores/sangue , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Materiais de Construção/análise , Endotélio Vascular/fisiopatologia , Humanos , Exposição por Inalação/análise , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Camundongos Endogâmicos C57BL , Cavidade Nasal/efeitos dos fármacos , Cavidade Nasal/imunologia , Cidade de Nova Iorque , Ataques Terroristas de 11 de Setembro , Células THP-1RESUMO
In order to ascertain the regulatory mechanism of fruit development in Isatis indigotica Fortune, the complementary DNA (cDNA) sequence of the SHATTERPROOF 2 (SHP2) orthologous gene was identified by Rapid Amplification of cDNA Ends technology and the corresponding gene was named IiSHP2. The expression pattern of IiSHP2 was determined by quantitative reverse transcription-polymerase chain reaction and wild-type Col-0 Arabidopsis plants were transformed with the IiSHP2 gene using Agrobacterium tumefaciens and the floral-dip method. Expression analyses indicated that IiSHP2 was highly expressed in flowers, silicles and seeds. Compared to wild-type plants, IiSHP2 transgenic lines bolted earlier. Detailed phenotypic observations showed that the size of the rosette and cauline leaves in transgenic lines was reduced and the cauline leaves of the transgenic lines were incurved and displayed a funnel-like shape. During the reproductive growth stage, IiSHP2 transgenic plants produced shortened sepals and the flower buds were not encapsulated completely. Moreover, the petals of the transgenic lines were converted into stamineous tissues, accompanied by exposed stamens, short malformed siliques and wrinkled valves, indicating a severe decline in fertility. These experimental conclusions are valuable as a reference for the breeding of medicinal plants.
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The circadian clock in plants synchronizes biological processes that display cyclic 24-h oscillation based on metabolic and physiological reactions. This clock is a precise timekeeping system, that helps anticipate diurnal changes; e.g., expression levels of clock-related genes move in synchrony with changes in pathogen infection and help prepare appropriate defense responses in advance. Salicylic acid (SA) is a plant hormone and immune signal involved in systemic acquired resistance (SAR)-mediated defense responses. SA signaling induces cellular redox changes, and degradation and rhythmic nuclear translocation of the non-expresser of PR genes 1 (NPR1) protein. Recent studies demonstrate the ability of the circadian clock to predict various potential attackers, and of redox signaling to determine appropriate defense against pathogen infection. Interaction of the circadian clock with redox rhythm promotes the balance between immunity and growth. We review here a variety of recent evidence for the intricate relationship between circadian clock and plant immune response, with a focus on the roles of redox rhythm and NPR1 in the circadian clock and plant immunity.
Assuntos
Relógios Circadianos , Imunidade Vegetal , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Ácido Salicílico/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
BACKGROUND: Myelination, degeneration and regeneration are implicated in crucial responses to injury in the peripheral nervous system. Considering the progression of amyotrophic lateral sclerosis (ALS), we used the superoxide dismutase 1 (SOD1)-G93A transgenic mouse model of ALS to investigate the effects of mutant SOD1 on the peripheral nerves. METHODS: Changes in peripheral nerve morphology were analyzed in SOD1 mutant mice at various stages of the disease by toluidine blue staining and electron microscopy (EM). Schwann cell proliferation and recruitment of inflammatory factors were detected by immunofluorescence staining and quantitative reverse transcription PCR and were compared between SOD1 mutant mice and control mice. Furthermore, western blotting (WB) and TUNEL staining were used to investigate axonal damage and Schwann cell survival in the sciatic nerves of mice in both groups. RESULTS: An analysis of the peripheral nervous system in SOD1-G93A mice revealed the following novel features: (i) Schwann cells and axons in mutant mice underwent changes that were similar to those seen in the control mice during the early development of peripheral nerves. (ii) The peripheral nerves of SOD1-G93A mice developed progressive neuropathy, which presented as defects in axons and myelin, leading to difficulty in walking and reduced locomotor capacity at a late stage of the disease. (iii) Macrophages were recruited and accumulated, and nerve injury and a deficit in the blood-nerve barrier were observed. (iv) Proliferation and the inflammatory micro-environment were inhibited, which impaired the regeneration and remyelination of axons after crush injury in the SOD1-G93A mice. CONCLUSIONS: The mutant human SOD1 protein induced axonal and myelin degeneration during the progression of ALS and participated in axon remyelination and regeneration in response to injury.
Assuntos
Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/patologia , Axônios/patologia , Nervos Periféricos/patologia , Células de Schwann/patologia , Superóxido Dismutase-1/genética , Superóxido Dismutase/genética , Esclerose Lateral Amiotrófica/fisiopatologia , Animais , Axônios/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Humanos , Camundongos , Camundongos Transgênicos , Regeneração Nervosa , Nervos Periféricos/metabolismo , Nervos Periféricos/fisiopatologia , Mutação Puntual , Células de Schwann/metabolismoRESUMO
To investigate the hepatoprotective effect of Schisandrae Chinensis Fructus (SCF) on CCl4-induced liver injury, observe its effect on serum metabolites, explore its scientific connotation in liver preservation and find the biomarkers for hepatoprotective effect of SCF. Liver injury model was established by using CCl4. The pathological sections of liver tissues were observed and the contents of alanine transaminase (ALT) and aspartate transaminase (AST) in serum were determined. The metabolic skills were adopted based on ultra performance liquid chromatography time-of-flight mass spectrometry (UPLC-Q-TOF-MS), principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) for screening and identification of biomarkers related to liver injury. The results showed the metabolites in blank group, model group and administration group could be easily distinguished, 50 differential compounds were identified and 7 possible metabolic pathways of liver protection were enriched. In this experiment, the hepatoprotective effect of SCF was verified, and the related metabolic pathways such as amino acid metabolism, vitamin metabolism and glycerophospholipid metabolism were discussed.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Medicamentos de Ervas Chinesas/química , Metabolômica , Schisandra/química , Animais , Biomarcadores , Cromatografia Líquida de Alta Pressão , Frutas/química , Fígado/efeitos dos fármacos , Fígado/metabolismo , Análise de Componente Principal , Espectrometria de Massas em TandemRESUMO
Fifteen SPL (SQUAMOSA PROMOTER BINDING PROTEIN-LIKE) genes were identified and characterized in Nicotiana tabacum L. cv. Qinyan95. The exon-intron structures of these genes were determined according to the coding sequences confirmed by RT-PCR and the genomic DNA sequences downloaded from the databases in Sol Genomics Network, and thirteen of them were found to carry the response element of miR156. To elucidate the origin of the validated NtabSPL genes, multiple alignments of the nucleotide sequences encompassing the open reading frames were conducted by using the orthologs in N. tabacum, Nicotiana sylvestris, Nicotiana tomentosiformis, and Nicotiana otophora. The results showed that six NtabSPL genes were derived from a progenitor of N. sylvestris, and nine NtabSPL genes were derived from a progenitor of N. tomentosiformis, further corroborating that N. tabacum came from the interspecific hybridization between the ancestors of N. sylvestris and N. tomentosiformis. In contrast to previous statements about highly repetitive sequences, the genome of N. tabacum mainly retained the paternal-derived SPL genes in diploidization process. Phylogenetic analyses based on the highly conserved SBP (SQUAMOSA PROMOTER BINDING PROTEIN) domains and the full-length amino acid sequences reveal that the SPL proteins of tobacco, tomato, and Arabidopsis can be categorized into eight groups. It is worth noting that N. tabacum contains seven NtabSPL6 genes originated from two parental genomes and NtabSPL6-2 possesses a GC-AG intron. In addition, transgenic tobacco plants harboring Arabidopsis Pri-miR156A were generated by Agrobacterium-mediated transformation method, and the constitutive expression of miR156 could obviously inhibit the activity of the NtabSPL genes containing its target site, suggesting the function of miR156 is conservative in tobacco and Arabidopsis.
Assuntos
Nicotiana/genética , Filogenia , Proteínas de Plantas/genética , Sequência de Bases , Evolução Molecular , Íntrons , Proteínas de Plantas/química , Alinhamento de Sequência , Nicotiana/classificaçãoRESUMO
Chlorpyrifos (CPF), dichlorvos (DDV), and cypermethrin (CP), as commonly used pesticides, have been implicated in inducing neuropsychiatric disorders, such as anxiety, depression-like behaviors, and locomotor activity impairment. However, the exact molecular mechanisms of these adverse effects, particularly in both sexes and their next-generation effects, remain unclear. In this study, we conducted behavioral analysis, along with cellular assays (monodansylcadaverine staining) and molecular investigations (qRT-PCR and western blotting of mTOR, P62, and Beclin-1) to clear the potential role of autophagy in pesticide-induced behavioral alterations. For this purpose, 42 adult female and 21 male inbred ICR mice (F0) were distributed into seven groups. Maternal mice (F0) and 112 F1 offspring were exposed to 0.5 and 1 ppm of CPF, DDV, and CP through drinking water. F1 male and female animals were studied to assess the sex-specific effects of pesticides on brain tissue. Our findings revealed pronounced anxiogenic effects and impaired locomotor activity in mice. F1 males exposed to CPF (1 ppm) exhibited significantly elevated depression-like behaviors compared to other groups. Moreover, pesticide exposure reduced mTOR and P62 levels, while enhancing the Beclin-1 gene and protein expression. These changes in autophagy signaling pathways, coupled with oxidative and neurogenic damage in the cerebral cortex and hippocampus, potentially contribute to heightened locomotor activity, anxiety, and depression-like behaviors following pesticide exposure. This study underscores the substantial impact of pesticides on both physiological and behavioral aspects, emphasizing the necessity for comprehensive assessments and regulatory considerations for pesticide use. Additionally, the identification of sex-specific responses presents a crucial dimension for pharmaceutical sciences, highlighting the need for tailored therapeutic interventions and further research in this field.
Assuntos
Ansiedade , Autofagia , Comportamento Animal , Depressão , Camundongos Endogâmicos ICR , Estresse Oxidativo , Praguicidas , Animais , Feminino , Masculino , Camundongos , Autofagia/efeitos dos fármacos , Ansiedade/induzido quimicamente , Ansiedade/fisiopatologia , Ansiedade/metabolismo , Depressão/metabolismo , Depressão/genética , Depressão/induzido quimicamente , Depressão/fisiopatologia , Estresse Oxidativo/efeitos dos fármacos , Praguicidas/toxicidade , Praguicidas/efeitos adversos , Comportamento Animal/efeitos dos fármacos , Locomoção/efeitos dos fármacos , Humanos , Serina-Treonina Quinases TOR/metabolismo , Serina-Treonina Quinases TOR/genética , Clorpirifos/toxicidade , Clorpirifos/efeitos adversosRESUMO
The AGL6 (AGMOUSE LIKE 6) gene is a member of the SEP subfamily and functions as an E-class floral homeotic gene in the development of floral organs. In this study, we cloned IiAGL6, the orthologous gene of AGL6 in Isatis indigotica. The constitutive expression of IiAGL6 in Arabidopsis thaliana resulted in a late-flowering phenotype and the development of curly leaves during the vegetative growth period. Abnormal changes in floral organ development were observed during the reproductive stage. In woad plants, suppression of IiAGL6 using TRV-VIGS (tobacco rattle virus-mediated virus-induced gene silencing) decreased the number of stamens and led to the formation of aberrant anthers. Similar changes in stamen development were also observed in miRNA-AGL6 transgenic Arabidopsis plants. Yeast two-hybrid and BiFC tests showed that IiAGL6 can interact with other MADS-box proteins in woad; thus, playing a key role in defining the identities of floral organs, particularly during stamen formation. These findings might provide novel insights and help investigate the biological roles of MADS transcription factors in I. indigotica.
Assuntos
Arabidopsis , Isatis , Isatis/genética , Isatis/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Domínio MADS/genética , Proteínas de Domínio MADS/metabolismo , Flores , Arabidopsis/metabolismo , Pólen/genética , Pólen/metabolismo , Regulação da Expressão Gênica de Plantas , Plantas Geneticamente Modificadas/metabolismo , FilogeniaRESUMO
Cellulose is an economical, biodegradable, widely available, and eco-friendly natural macromolecule. But its utilization has been restricted due to its insolubility in water and common organic solvents. In this work, soluble fluorescent probes based on cellulose were synthesized. Firstly, the primary hydroxyl group in glucose units was reacted with SOCl2 to introduce Cl and obtain chloro-cellulose (Cell-Cl). This operation breaks down the regular structure and hydrogen bonding of the original cellulose, enabling it to dissolve in DMSO. Secondly, the Cell-Cl reacted with CS2 and 2-mercaptobenzothiazole to obtain a cellulose-based macromolecular RAFT reagent (Cell-CTA). Finally, the fluorescent monomers which bears -C=C- and naphthalimide, and methacrylic acid (MAA) were grafted onto the main chain of cellulose through RAFT polymerization. Thus, cellulose-based readily soluble macromolecular fluorescent probes were obtained. The cellulose-based probes can specifically recognize Fe3+ in pure water and can be recycled and regenerated. Additionally, the cellulose-based probes exhibit remarkable adsorption and separation properties for Fe3+ ions. The modification of cellulose decreases its crystallinity and introduces hydrophilic groups and fluorophores, which enables cellulose to be soluble in both pure water and the organic solvent DMSO. This work expands the application range of cellulose-based copolymers.
Assuntos
Celulose , Corantes Fluorescentes , Celulose/química , Dimetil Sulfóxido , Polímeros/química , Solventes , ÁguaRESUMO
Green credit policy is the primary means for financial institutions to fulfill their environmental responsibilities. It is an issue worthy of attention whether green credit policy can achieve the effect of energy conservation, efficiency improvement, pollution reduction, and carbon reduction. This study uses the difference-in-difference method to test the impact of green credit policy on energy efficiency. The results show that green credit policy led to a significant decrease in energy intensity of green credit-restricted sectors while impeding the advancement of green total factor energy efficiency. The heterogeneity results show that the energy efficiency of large-scale, light textile manufacturing, resource processing industries, and clean industries are more significantly affected. Green credit policy can achieve energy conservation and has a linkage effect on pollution and carbon reduction. Although the constraint effect of green credit policy has effectively suppressed energy intensity, it also leads some industries to face a vicious cycle of "enhanced financing constraints-weakened innovation impetus," which in turn makes it challenging to improve green total factor energy efficiency. The above findings confirm the effectiveness of green credit policy in energy conservation and emission reduction. Also, they indicate the necessity of further improvement of the green financial policy system.
Assuntos
Conservação de Recursos Energéticos , Política Fiscal , Políticas , China , Carbono , Desenvolvimento EconômicoRESUMO
In this paper, a multilayer ultra-wideband transparent metamaterial wave absorber is proposed, which has the characteristics of ultra-wideband wave absorption, light transmission and flexible bending; in addition, due to the complete symmetry of the structure, the absorber has polarization insensitivity to incident electromagnetic waves. Both simulation and experimental results show that the frequency range of the microwave absorption rate is higher than 90% between 8.7 GHz and 38.9 GHz (between which most of the absorption rate can reach more than 95%), the total bandwidth is 30.2 GHz, and the relative bandwidth is 126.9%, realizing microwave broadband absorption and covering commonly used communication frequency bands such as X-band, Ku-band, and K-band. A sample was processed and tested. The test results are in good agreement with the results of the theoretical analysis, which proves the correctness of the theoretical analysis. In addition, through the selection and oxidation of indium tin (ITO) materials, the metamaterial also has the characteristics of optical transparency and flexibility, so it has potential application value in the window radar stealth and conformal radar stealth of weapons and equipment.
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Arsenic contamination in drinking water causes a global public health problem. Emerging evidence suggests that arsenic may act as an environmental risk factor for anxiety disorders. However, the exact mechanism underlying the adverse effects has not been fully elucidated. This study aimed to evaluate the anxiety-like behaviors of mice exposed to arsenic trioxide (As2O3), to observe the neuropathological changes, and to explore the link between the GABAergic system and behavioral manifestations. For this purpose, male C57BL/6 mice were exposed to various doses of As2O3 (0, 0.15, 1.5, and 15 mg/L) through drinking water for 12 weeks. Anxiety-like behaviors were assessed using the open field test (OFT), light/dark choice test, and elevated zero maze (EZM). Neuronal injuries in the cerebral cortex and hippocampus were assessed by light microscopy with H&E and Nissl staining. Ultrastructural alteration in the cerebral cortex was assessed by transmission electron microscope (TEM). The expression levels of GABAergic system-related molecules (i.e., glutamate decarboxylase, GABA transporter, and GABAB receptor subunits) in the prefrontal cortex (PFC) were determined by qRT-PCR and western blotting. Arsenic exposure showed a striking anxiogenic effect on mice, especially in the group exposed to 15 mg/L As2O3. Light microscopy showed neuron necrosis and reduced cell counts. TEM revealed marked ultrastructural changes, including the vacuolated mitochondria, disrupted Nissl bodies, an indentation in the nucleus membrane, and delamination of myelin sheath in the cortex. In addition, As2O3 influenced the GABAergic system in the PFC by decreasing the expression of the glutamate decarboxylase 1 (GAD1) and the GABAB2 receptor subunit, but not the GABAB1 receptor subunit. To sum up, sub-chronic exposure to As2O3 is associated with increased anxiety-like behaviors, which may be mediated by altered GABAergic signaling in the PFC. These findings shed light on the mechanisms responsible for the neurotoxic effects of arsenic and therefore more cautions should be taken.
Assuntos
Arsênio , Água Potável , Masculino , Camundongos , Animais , Arsênio/toxicidade , Camundongos Endogâmicos C57BL , Córtex Pré-Frontal/metabolismo , Córtex Pré-Frontal/patologia , Ansiedade/induzido quimicamente , Ácido gama-Aminobutírico/metabolismoRESUMO
Lead (Pb) is a highly toxic heavy metal. Pb exposure could adversely affect many organs, including the male reproductive system. Oxidative stress and mitochondrial impairment play a fundamental role in the pathogenesis of Pb-induced male reproductive system injury. Taurine (TAU) is abundantly found in mammalian bodies. The positive effects of TAU on oxidative stress biomarkers and mitochondrial function have been reported. The current study evaluated the effects of TAU on Pb-induced reproductive toxicity. Mice received Pb (20 mg/kg/day; gavage, 35 consecutive days). Then, sperm indices (quality and quantity) together with sperm kinetics, sperm mitochondrial parameters, testicular and sperm oxidative stress biomarkers, testis and plasma testosterone levels, and the expression of genes involved in the steroidogenesis process have been evaluated. Pb caused significant histopathological alterations and oxidative stress in male mice's reproductive system and sperm. Moreover, significant mitochondrial function impairment was evident in sperm isolated from Pb-treated mice. Pb exposure also suppressed the expression of StAR, 17ß-HSD, CYP11A, and 3ß-HSD genes in the male gonad. It was found that TAU (500 and 1000 mg/kg) significantly improved oxidative stress biomarkers in both male gonads and gametes of Pb-treated mice. TAU also significantly restored sperm mitochondrial function and kinetics. The expression of genes involved in steroidogenesis was also higher in TAU-treated animals. These data suggest TAU as an effective agent against Pb-induced reproductive toxicity. The effects of TAU on oxidative stress markers, mitochondrial function, and the steroidogenesis process seem to play a fundamental role in its protective properties. Further studies are warranted to detect the precise protective effects of this amino acid in the reproductive system. Lead (Pb) is a toxic element that adversely affects the male reproductive system. Mitochondrial impairment and oxidative stress have a crucial role in the Pb-induced reproductive toxicity. Taurine (TAU) could considerably improve the reproductive toxicity induced by Pb via enhancing mitochondrial function and mitigating oxidative stress indices. ΔΨ, mitochondrial membrane potential; ATP, adenosine triphosphate.
Assuntos
Chumbo , Taurina , Masculino , Camundongos , Animais , Taurina/farmacologia , Taurina/metabolismo , Fenômenos Biomecânicos , Chumbo/toxicidade , Chumbo/metabolismo , Sêmen/metabolismo , Espermatozoides/metabolismo , Testículo/metabolismo , Estresse Oxidativo , Mitocôndrias/metabolismo , Biomarcadores/metabolismo , Testosterona , Mamíferos/metabolismoRESUMO
Epidemiological and experimental studies have demonstrated the atherogenic effects of environmental toxicant arsenic and fluoride. Inflammatory mechanism plays an important role in the pathogenesis of atherosclerosis. The aim of the present study is to determine the effect of chronic exposure to arsenic and fluoride alone or combined on inflammatory response in rabbit aorta. We analyzed the expression of genes involved in leukocyte adhesion [P-selectin (P-sel) and vascular cell adhesion molecule-1(VCAM-1)], recruitment and transendothelial migration of leukocyte [interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1)] and those involved in pro-inflammatory cytokines [interleukin-6 (IL-6)]. We found that fluoride and arsenic alone or combined increased the expression of VCAM-1, P-sel, MCP-1, IL-8, and IL-6 at the RNA and protein levels. The gene expressions of inflammatory-related molecules were attenuated when co-exposure to the two toxicants compared with just one of them. We also examined the lipid profile of rabbits exposed to fluoride and (or) arsenic. The results showed that fluoride slightly increased the serum lipids but arsenic decreased serum triglyceride. We showed that inflammatory responses but not lipid metabolic disorder may play a crucial role in the mechanism of the cardiovascular toxicity of arsenic and fluoride.
Assuntos
Aorta/efeitos dos fármacos , Arsênio/toxicidade , Moléculas de Adesão Celular/efeitos dos fármacos , Moléculas de Adesão Celular/genética , Fluoretos/toxicidade , Vasculite/induzido quimicamente , Animais , Aorta/metabolismo , Exposição Ambiental , Feminino , Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica , Fenômenos do Sistema Imunitário , Leucócitos , Masculino , Coelhos , Testes de Toxicidade Crônica , Molécula 1 de Adesão de Célula Vascular/efeitos dos fármacos , Molécula 1 de Adesão de Célula Vascular/genética , Vasculite/imunologiaRESUMO
Hyaluronic acid (HA), a high-value biomacromolecule, has wide applications in medical, cosmetic and food fields. Currently, employing the safe-grade microorganisms for de novo biosynthesis of HA from renewable substrates has become a promising alternative. In this study, we established a Bacillus amyloliquefaciens strain as platform for HA production from Jerusalem artichoke inulin. Firstly, the different HA and UDP-GlcUA synthase genes were introduced into B. amyloliquefaciens to construct the HA synthesis pathway. Secondly, the byproduct polysaccharides were removed by knocking sacB and epsA-O using CRISPR/Cas9n system, resulting in a 13% increase in HA production. Finally, 2.89 g/L HA with a high molecular weight of 1.5 MDa was obtained after optimizing fermentation conditions and adding osmotic agents. This study demonstrates the engineered B. amyloliquefaciens can effectively synthesize HA with Jerusalem artichoke inulin and provides a green route for HA production.
Assuntos
Bacillus amyloliquefaciens , Helianthus , Bacillus amyloliquefaciens/genética , Bacillus amyloliquefaciens/metabolismo , Fermentação , Helianthus/genética , Helianthus/metabolismo , Ácido Hialurônico/metabolismo , Inulina/metabolismoRESUMO
Ectoine, an osmotic pressure-compensated solute, is used in the food, agriculture, medicine, and cosmetics industries due to its ability to protect macromolecules. In this study, an ectoine-producing variant of Escherichia coli, ET08, was genetically constructed by introducing the ectABC gene cluster and eliminating metabolic pathways involving lysine and pyruvate. Medium optimization enhanced ectoine production from 1.87 to 10.2 g/L. Analysis of the transcriptional levels revealed that supplementation with ammonium sulfate enhanced the metabolic flux towards the biosynthesis of ectoine. Furthermore, by optimizing the copy number of ectA, ectB, and ectC, the recombinant E. coli ET11 (ectA:ectB:ectC = 1:2:1) produced 12.9 g/L ectoine in the shake flask and 53.2 g/L ectoine in a fed-batch fermenter, representing the highest ectoine titer produced by E. coli, which has great industrial prospects.
RESUMO
Taurine (TAU) is a free amino acid abundant in the human body. Various physiological roles have been attributed to TAU. At the subcellular level, mitochondria are the primary targets for TAU function. Meanwhile, it has been found that TAU depletion is associated with severe pathologies. Cholestasis is a severe clinical complication that can progress to liver fibrosis, cirrhosis, and hepatic failure. Bile duct ligation (BDL) is a reliable model for assessing cholestasis/cirrhosis and related complications. The current study was designed to investigate the effects of cholestasis/cirrhosis on tissue and mitochondrial TAU reservoirs. Cholestatic rats were monitored (14 and 42 days after BDL surgery), and TAU levels were assessed in various tissues and isolated mitochondria. There was a significant decrease in TAU in the brain, heart, liver, kidney, skeletal muscle, intestine, lung, testis, and ovary of the BDL animals (14 and 42 days after surgery). Mitochondrial levels of TAU were also significantly depleted in BDL animals. Tissue and mitochondrial TAU levels in cirrhotic animals (42 days after the BDL operation) were substantially lower than those in the cholestatic rats (14 days after BDL surgery). These data indicate an essential role for tissue and mitochondrial TAU in preventing organ injury induced by cholestasis/cirrhosis and could justify TAU supplementation for therapeutic purposes.