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1.
Appl Opt ; 63(4): 874-887, 2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-38437383

RESUMO

A single Raman cell configuration useful for DIAL ozone lidar is designed and optimized. The conversion efficiency and flexibility of using a single Raman cell filled with a mixture of high pressure Raman active gases hydrogen (H 2) and methane (C H 4) have been examined and reported. The stimulated Raman scattering (SRS) conversion efficiency of Raman active gases with different total cell pressures and the volume mixing ratio excited with a focused, frequency quadrupled Nd:YAG laser with a maximum pulse energy of 25 mJ and a pulse duration of 10 ns at 100 Hz repetition rate are examined in detail. The gas combination of H 2:C H 4 emits a coaxial beam of two wavelengths, 288.4 nm (C H 4) and 299.1 nm (H 2), with a maximum total conversion efficiency of about 45%. The optimum volume mixing ratio for generating the required wavelength pair with almost equal energies is found to be 2:1 (H 2:C H 4) at a total cell pressure of 18 bar. The contribution of cascade Raman scattering (CRS) and four-wave mixing (FWM) to the higher order Stokes lines is examined. The laser attenuation due to soot formation under various mixing ratios in the cell is also presented.

2.
PLoS Biol ; 18(3): e3000470, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32150534

RESUMO

In the spinal cord, the central canal forms through a poorly understood process termed dorsal collapse that involves attrition and remodelling of pseudostratified ventricular layer (VL) cells. Here, we use mouse and chick models to show that dorsal ventricular layer (dVL) cells adjacent to dorsal midline Nestin(+) radial glia (dmNes+RG) down-regulate apical polarity proteins, including Crumbs2 (CRB2) and delaminate in a stepwise manner; live imaging shows that as one cell delaminates, the next cell ratchets up, the dmNes+RG endfoot ratchets down, and the process repeats. We show that dmNes+RG secrete a factor that promotes loss of cell polarity and delamination. This activity is mimicked by a secreted variant of Crumbs2 (CRB2S) which is specifically expressed by dmNes+RG. In cultured MDCK cells, CRB2S associates with apical membranes and decreases cell cohesion. Analysis of Crb2F/F/Nestin-Cre+/- mice, and targeted reduction of Crb2/CRB2S in slice cultures reveal essential roles for transmembrane CRB2 (CRB2TM) and CRB2S on VL cells and dmNes+RG, respectively. We propose a model in which a CRB2S-CRB2TM interaction promotes the progressive attrition of the dVL without loss of overall VL integrity. This novel mechanism may operate more widely to promote orderly progenitor delamination.


Assuntos
Proteínas de Membrana/metabolismo , Medula Espinal/citologia , Medula Espinal/embriologia , Animais , Adesão Celular , Embrião de Galinha , Cães , Regulação da Expressão Gênica no Desenvolvimento , Células HEK293 , Humanos , Células Madin Darby de Rim Canino , Proteínas de Membrana/genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Junções Íntimas/metabolismo , Imagem com Lapso de Tempo
3.
J Basic Microbiol ; 62(2): 185-196, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34913505

RESUMO

Bacterial blight (BB) of rice is a devastating disease caused by Xanthomonas oryzae pv. oryzae (Xoo). The evolution of new pathogenic races of bacterial blight pathogen is always a potential threat for rice production. The deployment of pathotype-specific resistant genes in the host plants is a feasible strategy to develop BB-resistant varieties. Therefore, continuous disease monitoring, identification of Xoo pathotypes, and their distribution are crucial to managing BB. In this study, 71 Xoo isolates were collected from the Godavari delta in Andhra Pradesh (India) and their virulence profiles on rice BB differentials were characterized. Data revealed that different International Rice Bacterial Blight (IRBB) lines with single BB resistance genes were susceptible to 73.2%-97.2% of the isolates, except IRBB13 (possessing BB resistance gene, xa13) which showed a moderately susceptible or susceptible reaction to 47.9% of the isolates. Three gene combination rice differentials like IRBB56 (Xa4 + xa5 + xa13), IRBB57 (Xa4 + xa5 + Xa21), IRBB58 (Xa4 + xa13 + Xa21), and IRBB59 (xa5 + xa13 + Xa21) showed very broad-spectrum resistance to majority of the Xoo isolates from the region. None of the tested Xoo isolates were virulent on IRBB58 (Xa4 + xa13 + Xa21), IRBB60 (Xa4 + xa5 + xa13 + Xa21), and IRBB66 (Xa4 + xa5 + Xa7 + xa13 + Xa21). Based on the virulence reaction, 71 Xoo isolates were grouped into 10 major pathotypes. Highly virulent pathotypes viz., IXoPt # 14, 17, 19, and 22 can break the resistance of major BB-resistant genes and were commonly distributed throughout the surveyed regions. Genotypic data of 71 Xoo isolates using J3 primer divided them into three major clusters. Cluster I consisted of 24 Xoo isolates that belonged to pathotype IXoPt-19. Cluster II consisted of 41 Xoo isolates belonging to seven different pathotypes, and Cluster III was composed of six isolates from three different pathotypes. The findings of this study will be helpful to develop rice varieties with pathotype-specific broad-spectrum resistance against BB.


Assuntos
Oryza , Xanthomonas , Genótipo , Doenças das Plantas , Xanthomonas/genética
4.
Proc Natl Acad Sci U S A ; 112(38): E5343-50, 2015 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-26354122

RESUMO

The phylum Apicomplexa includes serious pathogens of humans and animals. Understanding the distribution and population structure of these protozoan parasites is of fundamental importance to explain disease epidemiology and develop sustainable controls. Predicting the likely efficacy and longevity of subunit vaccines in field populations relies on knowledge of relevant preexisting antigenic diversity, population structure, the likelihood of coinfection by genetically distinct strains, and the efficiency of cross-fertilization. All four of these factors have been investigated for Plasmodium species parasites, revealing both clonal and panmictic population structures with exceptional polymorphism associated with immunoprotective antigens such as apical membrane antigen 1 (AMA1). For the coccidian Toxoplasma gondii only genomic diversity and population structure have been defined in depth so far; for the closely related Eimeria species, all four variables are currently unknown. Using Eimeria tenella, a major cause of the enteric disease coccidiosis, which exerts a profound effect on chicken productivity and welfare, we determined population structure, genotype distribution, and likelihood of cross-fertilization during coinfection and also investigated the extent of naturally occurring antigenic diversity for the E. tenella AMA1 homolog. Using genome-wide Sequenom SNP-based haplotyping, targeted sequencing, and single-cell genotyping, we show that in this coccidian the functionality of EtAMA1 appears to outweigh immune evasion. This result is in direct contrast to the situation in Plasmodium and most likely is underpinned by the biology of the direct and acute coccidian life cycle in the definitive host.


Assuntos
Variação Antigênica , Eimeria tenella/genética , Eimeria tenella/imunologia , Animais , Antígenos de Protozoários/imunologia , Sequência de Bases , Galinhas/parasitologia , Coccidiose/parasitologia , Cruzamentos Genéticos , Fezes , Variação Genética , Genética Populacional , Genótipo , Geografia , Dados de Sequência Molecular , Oocistos , Filogenia , Plasmodium/genética , Plasmodium/imunologia , Polimorfismo de Nucleotídeo Único , Doenças das Aves Domésticas/parasitologia , Vacinas Protozoárias
5.
Development ; 141(16): 3266-76, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25063452

RESUMO

Here, we exploit the spatial separation of temporal events of neural differentiation in the elongating chick body axis to provide the first analysis of transcriptome change in progressively more differentiated neural cell populations in vivo. Microarray data, validated against direct RNA sequencing, identified: (1) a gene cohort characteristic of the multi-potent stem zone epiblast, which contains neuro-mesodermal progenitors that progressively generate the spinal cord; (2) a major transcriptome re-organisation as cells then adopt a neural fate; and (3) increasing diversity as neural patterning and neuron production begin. Focussing on the transition from multi-potent to neural state cells, we capture changes in major signalling pathways, uncover novel Wnt and Notch signalling dynamics, and implicate new pathways (mevalonate pathway/steroid biogenesis and TGFß). This analysis further predicts changes in cellular processes, cell cycle, RNA-processing and protein turnover as cells acquire neural fate. We show that these changes are conserved across species and provide biological evidence for reduced proteasome efficiency and a novel lengthening of S phase. This latter step may provide time for epigenetic events to mediate large-scale transcriptome re-organisation; consistent with this, we uncover simultaneous downregulation of major chromatin modifiers as the neural programme is established. We further demonstrate that transcription of one such gene, HDAC1, is dependent on FGF signalling, making a novel link between signals that control neural differentiation and transcription of a core regulator of chromatin organisation. Our work implicates new signalling pathways and dynamics, cellular processes and epigenetic modifiers in neural differentiation in vivo, identifying multiple new potential cellular and molecular mechanisms that direct differentiation.


Assuntos
Cromatina/metabolismo , Neurogênese/fisiologia , Neurônios/citologia , Transcriptoma , Animais , Padronização Corporal , Ciclo Celular , Diferenciação Celular , Linhagem da Célula , Embrião de Galinha , Epigênese Genética , Fatores de Crescimento de Fibroblastos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Histona Desacetilase 1/metabolismo , Camundongos , Análise de Sequência com Séries de Oligonucleotídeos , Análise de Sequência de RNA , Transdução de Sinais , Medula Espinal/embriologia , Fatores de Tempo , Fator de Crescimento Transformador beta/metabolismo
6.
J Eur Acad Dermatol Venereol ; 31(8): 1324-1332, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28556418

RESUMO

BACKGROUND: OX40 (CD134) is expressed in lesional but not healthy skin of patients with psoriasis. KHK4083 is a fully human monoclonal antibody against OX40. OBJECTIVE: The primary aim of this first-in-human phase 1 study was to determine the safety and tolerability of ascending single doses of KHK4083 in patients with mild to moderate plaque psoriasis. Secondary aims were to determine the pharmacokinetics and immunogenicity of KHK4083, and an exploratory objective was to assess clinical activity. METHODS: In phase 1a, single doses of KHK4083 0.003 and 0.001 mg/kg IV were administered open label in two cohorts (each n = 6). Phase 1b had a multicentre, randomized, double-blind, placebo-controlled, ascending single-dose design in seven cohorts. Randomization was performed 3 : 1 to KHK4083 (n = 6) or placebo (n = 2) within each cohort. Ascending doses of KHK4083 were 0.03, 0.1, 0.3, 1.0, 3.0 and 10 mg/kg IV, and 1.0 mg/kg SC. RESULTS: There were no severe or serious adverse events (AEs), or discontinuations because of AEs. The most frequent treatment-related AEs in the 55 patients who received KHK4083 were mild or moderate chills (9.1%), and infusion/injection site reactions (7.3%). No clinically meaningful or dose-related changes from baseline in laboratory values, vital signs, ECG recordings or physical examinations were observed. Some KHK4083 recipients (10/54) developed anti-KHK4083 antibodies following treatment. Mean elimination half-life (t1/2 ) increased with dose, maximum serum concentration increased in a dose-proportional manner, and area under the serum concentration-time curve increased in a more than dose-proportional manner with increasing IV dose. Absolute bioavailability following SC administration was 73%. There was some indication of improvement in Psoriasis Area Severity Index (PASI) and sPGA scores at the highest IV doses (1.0 and 10 mg/kg) and the SC dose (1.0 mg/kg). The largest PASI 50 response and improvement in sPGA score ≥2 occurred with KHK4083 1.0 mg/kg SC. CONCLUSION: KHK4083 administration as a single dose up to 10 mg/kg IV or 1.0 mg/kg SC was generally safe and well tolerated in patients with mild to moderate plaque psoriasis with no dose-limiting AEs.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Psoríase/tratamento farmacológico , Receptores OX40/antagonistas & inibidores , Receptores OX40/imunologia , Adulto , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/efeitos adversos , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais Humanizados , Estudos de Coortes , Relação Dose-Resposta a Droga , Feminino , Humanos , Reação no Local da Injeção , Injeções Subcutâneas , Masculino , Pessoa de Meia-Idade , Placebos , Índice de Gravidade de Doença , Resultado do Tratamento
7.
Br J Dermatol ; 174(6): 1266-76, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26800231

RESUMO

BACKGROUND: Plaque psoriasis is a chronic and often debilitating skin disorder and proinflammatory cytokines are known to play a key role in the disease process. OBJECTIVES: To evaluate the safety and efficacy of baricitinib, an oral Janus kinase (JAK) 1/JAK2 inhibitor, in patients with moderate-to-severe psoriasis in a randomized, double-blind, placebo-controlled, dose-ranging phase 2b study. METHODS: Patients were randomized (n = 271) to receive placebo or oral baricitinib at 2, 4, 8 or 10 mg once daily for 12 weeks (Part A). Dose adjustment for 12 additional weeks was based on percentage improvement in the Psoriasis Area and Severity Index (PASI) score. The primary end point was Psoriasis Area and Severity Index (PASI) 75% (PASI-75) at 12 weeks for North American patients (n = 238); secondary end points were safety and efficacy measures in the entire population. RESULTS: At week 12, more North American patients in the 8-mg (43%) and 10-mg (54%) baricitinib groups than in placebo group (17%; P < 0·05) achieved PASI-75. All baricitinib-treated groups had greater mean changes from baseline in their PASI scores (P < 0·05) at 12 weeks and (except 2 mg) had higher rates of PASI-50 than the placebo group; statistically significant PASI-90 responses were achieved in the 8-mg and 10-mg groups at 8 and 12 weeks. More than 81% of PASI-75 responders maintained their scores through 24 weeks. During Part A, study discontinuations due to adverse events (AEs) were 0%, 0%, 2·8%, 6·3% and 5·8% and treatment-emergent AE rates were 44%, 50%, 47%, 58% and 64% for placebo and 2-, 4-, 8- and 10-mg baricitinib groups, respectively. No opportunistic infections were observed in any treatment group. Dose-dependent changes in laboratory values were observed. CONCLUSIONS: Patients with moderate-to-severe psoriasis treated with baricitinib for 12 weeks achieved significant improvements in PASI-75.


Assuntos
Azetidinas/administração & dosagem , Fármacos Dermatológicos/administração & dosagem , Psoríase/tratamento farmacológico , Sulfonamidas/administração & dosagem , Azetidinas/efeitos adversos , Fármacos Dermatológicos/efeitos adversos , Relação Dose-Resposta a Droga , Método Duplo-Cego , Esquema de Medicação , Feminino , Humanos , Janus Quinase 1/antagonistas & inibidores , Janus Quinase 2/antagonistas & inibidores , Masculino , Pessoa de Meia-Idade , Purinas , Pirazóis , Sulfonamidas/efeitos adversos , Resultado do Tratamento
8.
Br J Dermatol ; 175(5): 902-911, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27423107

RESUMO

BACKGROUND: Despite unmet need, 15 years have passed since a topical therapy with a new mechanism of action for atopic dermatitis (AD) has been approved. Janus kinase (JAK) inhibitor treatment effect via topical application in patients with AD is unknown. OBJECTIVES: Tofacitinib, a small-molecule JAK inhibitor, was investigated for the topical treatment of AD. METHODS: In this 4-week, phase IIa, randomized, double-blind, vehicle-controlled study (NCT02001181), 69 adults with mild-to-moderate AD were randomized 1:1 to 2% tofacitinib or vehicle ointment twice daily. Percentage change from baseline (CFB) in Eczema Area and Severity Index (EASI) score at week 4 was the primary end point. Secondary efficacy end points included percentage CFB in body surface area (BSA), CFB in EASI Clinical Signs Severity Sum Score, proportion of patients with Physician's Global Assessment (PGA) response and CFB in patient-reported pruritus. Safety, local tolerability and pharmacokinetics were monitored. RESULTS: The mean percentage CFB at week 4 in EASI score was significantly greater (P < 0·001) for tofacitinib (-81·7%) vs. vehicle (-29·9%). Patients treated with tofacitinib showed significant (P < 0·001) improvements vs. vehicle across all prespecified efficacy end points and for pruritus at week 4. Significant improvements in EASI, PGA and BSA were observed by week 1 and improvements in pruritus were observed by day 2. Safety/local tolerability were generally similar for both treatments, although more adverse events were observed for vehicle vs. tofacitinib. CONCLUSIONS: Tofacitinib ointment showed significantly greater efficacy vs. vehicle across end points, with early onset of effect and comparable safety/local tolerability to vehicle. JAK inhibition through topical delivery is potentially a promising therapeutic target for AD.


Assuntos
Dermatite Atópica/tratamento farmacológico , Piperidinas/administração & dosagem , Inibidores de Proteínas Quinases/administração & dosagem , Pirimidinas/administração & dosagem , Pirróis/administração & dosagem , Adolescente , Adulto , Método Duplo-Cego , Esquema de Medicação , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pomadas , Piperidinas/efeitos adversos , Piperidinas/farmacocinética , Inibidores de Proteínas Quinases/efeitos adversos , Inibidores de Proteínas Quinases/farmacocinética , Pirimidinas/efeitos adversos , Pirimidinas/farmacocinética , Pirróis/efeitos adversos , Pirróis/farmacocinética , Resultado do Tratamento , Adulto Jovem
9.
J Med Syst ; 40(11): 239, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27665113

RESUMO

The impact of internet and information systems across various domains have resulted in substantial generation of multidimensional datasets. The use of data mining and knowledge discovery techniques to extract the original information contained in the multidimensional datasets play a significant role in the exploitation of complete benefit provided by them. The presence of large number of features in the high dimensional datasets incurs high computational cost in terms of computing power and time. Hence, feature selection technique has been commonly used to build robust machine learning models to select a subset of relevant features which projects the maximal information content of the original dataset. In this paper, a novel Rough Set based K - Helly feature selection technique (RSKHT) which hybridize Rough Set Theory (RST) and K - Helly property of hypergraph representation had been designed to identify the optimal feature subset or reduct for medical diagnostic applications. Experiments carried out using the medical datasets from the UCI repository proves the dominance of the RSKHT over other feature selection techniques with respect to the reduct size, classification accuracy and time complexity. The performance of the RSKHT had been validated using WEKA tool, which shows that RSKHT had been computationally attractive and flexible over massive datasets.


Assuntos
Inteligência Artificial , Mineração de Dados/métodos , Sistemas de Apoio a Decisões Clínicas/organização & administração , Humanos , Aprendizado de Máquina
10.
Trop Anim Health Prod ; 48(7): 1455-62, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27509876

RESUMO

Anthelmintic resistance (AR) status in Madras Red sheep from selected field flocks of a government funded scheme, covered by regular, sustained anthelmintic treatment for more than 10 years was determined. Parameters such as fecal egg count reduction test (FECRT), larval paralysis assay (LPA), and allele-specific-PCR (AS-PCR) were used to test the efficacy of fenbendazole, tetramisole, and ivermectin at recommended doses, in two seasons. Sheep belonging to non-beneficiary farmers were used as controls. Mean FECRT values of beneficiary group during winter and summer seasons were 77.77 and 76.04, 93.65 and 92.12, and 95.37 and 98.06 %, respectively, for fenbendazole, tetramisole, and ivermectin. In the non-beneficiary groups, the corresponding values were 74.82 and 81.09 %, 96.05 and 97.40 %, and 97.26 and 98.23 %, respectively. The results revealed resistance to fenbendazole, suspect resistance to tetramisole and susceptibility to ivermectin in beneficiary flock. In non-beneficiary flock, while resistance was noticed against fenbendazole, both tetramisole and ivermectin were effective. FECR values were found to be significantly different between beneficiary and non-beneficiary groups against tetramisole. The results of LPA confirmed this finding, as 50 % of the Haemonchus contortus larvae were paralyzed at the concentration of 0.0156 µg/ml in the beneficiary group, while those of non-beneficiary groups required lower concentrations of 0.0078 µg/ml. AS-PCR revealed the predominance of heterozygous susceptible population of H. contortus in the beneficiary group. In this study, resistance to fenbendazole was confirmed in both the beneficiary and non-beneficiary groups and this could be attributed to frequent use of benzimidazoles as seen from the deworming records. Emergence of tetramisole resistance was detected in the beneficiary group, where the drug was used continuously for 4 years. Ivermectin was found to be effective in all the flocks. It is recommended that the practice of routine deworming of three to four times a year should be avoided, as it can lead to emergence of anthelmintic resistance.


Assuntos
Anti-Helmínticos/uso terapêutico , Resistência a Medicamentos , Hemoncose/veterinária , Doenças dos Ovinos/tratamento farmacológico , Ovinos/fisiologia , Criação de Animais Domésticos , Animais , Anti-Helmínticos/administração & dosagem , Anti-Helmínticos/farmacologia , Esquema de Medicação , Fezes/parasitologia , Fenbendazol/administração & dosagem , Fenbendazol/farmacologia , Fenbendazol/uso terapêutico , Hemoncose/tratamento farmacológico , Haemonchus/efeitos dos fármacos , Índia , Ivermectina/administração & dosagem , Ivermectina/farmacologia , Ivermectina/uso terapêutico , Contagem de Ovos de Parasitas/veterinária , Tetramizol/administração & dosagem , Tetramizol/farmacologia , Tetramizol/uso terapêutico , Clima Tropical
11.
Development ; 138(10): 1893-902, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21521736

RESUMO

The size, composition and functioning of the spinal cord is likely to depend on appropriate numbers of progenitor and differentiated cells of a particular class, but little is known about how cell numbers are controlled in specific cell cohorts along the dorsoventral axis of the neural tube. Here, we show that FatJ cadherin, identified in a large-scale RNA interference (RNAi) screen of cadherin genes expressed in the neural tube, is localised to progenitors in intermediate regions of the neural tube. Loss of function of FatJ promotes an increase in dp4-vp1 progenitors and a concomitant increase in differentiated Lim1(+)/Lim2(+) neurons. Our studies reveal that FatJ mediates its action via the Hippo pathway mediator Yap1: loss of downstream Hippo components can rescue the defect caused by loss of FatJ. Together, our data demonstrate that RNAi screens are feasible in the chick embryonic neural tube, and show that FatJ acts through the Hippo pathway to regulate cell numbers in specific subsets of neural progenitor pools and their differentiated progeny.


Assuntos
Proteínas Aviárias/metabolismo , Caderinas/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Animais , Proteínas Aviárias/antagonistas & inibidores , Proteínas Aviárias/genética , Sequência de Bases , Caderinas/antagonistas & inibidores , Caderinas/genética , Contagem de Células , Embrião de Galinha , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Silenciamento de Genes , Tubo Neural/citologia , Tubo Neural/embriologia , Tubo Neural/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Interferência de RNA , RNA Interferente Pequeno/genética , Transdução de Sinais
12.
Plant Physiol ; 162(1): 9-23, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23463782

RESUMO

The plant cell wall has many significant structural and physiological roles, but the contributions of the various components to these roles remain unclear. Modification of cell wall properties can affect key agronomic traits such as disease resistance and plant growth. The plant cell wall is composed of diverse polysaccharides often decorated with methyl, acetyl, and feruloyl groups linked to the sugar subunits. In this study, we examined the effect of perturbing cell wall acetylation by making transgenic Arabidopsis (Arabidopsis thaliana) and Brachypodium (Brachypodium distachyon) plants expressing hemicellulose- and pectin-specific fungal acetylesterases. All transgenic plants carried highly expressed active Aspergillus nidulans acetylesterases localized to the apoplast and had significant reduction of cell wall acetylation compared with wild-type plants. Partial deacetylation of polysaccharides caused compensatory up-regulation of three known acetyltransferases and increased polysaccharide accessibility to glycosyl hydrolases. Transgenic plants showed increased resistance to the fungal pathogens Botrytis cinerea and Bipolaris sorokiniana but not to the bacterial pathogens Pseudomonas syringae and Xanthomonas oryzae. These results demonstrate a role, in both monocot and dicot plants, of hemicellulose and pectin acetylation in plant defense against fungal pathogens.


Assuntos
Acetilesterase/metabolismo , Arabidopsis/fisiologia , Aspergillus nidulans/enzimologia , Brachypodium/fisiologia , Parede Celular/metabolismo , Polissacarídeos/metabolismo , Acetilação , Acetilesterase/genética , Arabidopsis/citologia , Arabidopsis/genética , Arabidopsis/imunologia , Ascomicetos/patogenicidade , Aspergillus nidulans/genética , Botrytis/patogenicidade , Brachypodium/citologia , Brachypodium/genética , Brachypodium/imunologia , Resistência à Doença , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Regulação da Expressão Gênica de Plantas , Glucanos/metabolismo , Peróxido de Hidrogênio/metabolismo , Pectinas/metabolismo , Componentes Aéreos da Planta , Doenças das Plantas/imunologia , Plantas Geneticamente Modificadas , Pseudomonas syringae/patogenicidade , Regulação para Cima , Xanthomonas/patogenicidade
13.
EMBO Rep ; 13(5): 448-54, 2012 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-22491029

RESUMO

Inheritance of apical membrane is proposed to maintain vertebrate neural stem cell proliferation. However, evidence for this is contradictory. Using direct clonal analysis and live imaging in chick neural tube, we show that divisions that separate apical and basal components generate an apical daughter, which becomes a neuron, and a basal daughter, which rapidly re-establishes apico-basal polarity and divides again. Using a recently described real-time reporter of Notch activity, we confirm progenitor status and demonstrate that division orientation can influence Notch signalling. In addition, we reveal loss of apical complex proteins on neuronal differentiation onset, suggesting that removal of this inherited complex is part of the neuronal differentiation mechanism. These findings reconcile contradictory data, link asymmetric division to Notch signalling dynamics and identify apical complex loss as a new step towards neuronal differentiation.


Assuntos
Tubo Neural/metabolismo , Receptores Notch/metabolismo , Fuso Acromático/metabolismo , Animais , Diferenciação Celular/fisiologia , Embrião de Galinha , Galinhas , Transdução de Sinais/fisiologia
14.
Methods Mol Biol ; 2831: 315-324, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39134859

RESUMO

The cell intrinsic mechanisms directing peripheral nerve regeneration have remained largely understudied, thus limiting our understanding of these processes and constraining the advancement of novel clinical therapeutics. The use of primary adult rat dorsal root ganglion (DRG) neurons cultured in vitro is well established. Despite this, these cells can be challenging to culture and have so far not been amenable to robust transfection or live-cell imaging. The ability to transfect these cells with fluorescent plasmid constructs to label subcellular structures, combined with high resolution time-lapse imaging has the potential to provide invaluable insight into how peripheral neurons coordinate their regenerative response, and which specific cellular structures are involved in this process. Here we describe a protocol that facilitates transfection and subsequent live-imaging of adult rat DRG neurons.


Assuntos
Gânglios Espinais , Regeneração Nervosa , Neurônios , Animais , Gânglios Espinais/citologia , Regeneração Nervosa/fisiologia , Ratos , Neurônios/citologia , Neurônios/fisiologia , Neurônios/metabolismo , Células Cultivadas , Transfecção/métodos , Imagem com Lapso de Tempo/métodos
15.
Methods Mol Biol ; 2831: 97-111, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39134846

RESUMO

To investigate the cell behavior underlying neuronal differentiation in a physiologically relevant context, differentiating neurons must be studied in their native tissue environment. Here, we describe an accessible protocol for fluorescent live imaging of differentiating neurons within ex vivo embryonic chicken spinal cord slice cultures, which facilitates long-term observation of individual cells within developing tissue.


Assuntos
Diferenciação Celular , Eletroporação , Neurônios , Medula Espinal , Animais , Eletroporação/métodos , Medula Espinal/citologia , Medula Espinal/embriologia , Embrião de Galinha , Neurônios/citologia , Neurônios/metabolismo , Galinhas , Neurogênese
16.
Plant Physiol Biochem ; 213: 108836, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38941724

RESUMO

The root system architecture is an important complex trait in rice. With changing climatic conditions and soil nutrient deficiencies, there is an immediate need to breed nutrient-use-efficient rice varieties with robust root system architectural (RSA) traits. To map the genomic regions associated with crucial component traits of RSA viz. root length and root volume, a biparental F2 mapping population was developed using TI-128, an Ethyl Methane Sulphonate (EMS) mutant of a mega variety BPT-5204 having high root length (RL) and root volume (RV) with wild type BPT-5204. Extreme bulks having high RL and RV and low RL and RV were the whole genome re-sequenced along with parents. Genetic mapping using the MutMap QTL-Seq approach elucidated two genomic intervals on Chr.12 (3.14-3.74 Mb, 18.11-20.85 Mb), and on Chr.2 (23.18-23.68 Mb) as potential regions associated with both RL and RV. The Kompetitive Allele Specific PCR (KASP) assays for SNPs with delta SNP index near 1 were associated with higher RL and RV in the panel of sixty-two genotypes varying in root length and volume. The KASP_SNPs viz. Chr12_S4 (C→T; Chr12:3243938), located in the 3' UTR region of LOC_Os12g06670 encoding a protein kinase domain-containing protein and Chr2_S6 (C→T; Chr2:23181622) present upstream in the regulator of chromosomal condensation protein LOC_Os2g38350. Validation of these genes using qRT-PCR and in-silico studies using various online tools and databases revealed higher expression in TI-128 as compared to BPT- 5204 at the seedling and panicle initiation stages implying the functional role in enhancing RL and RV.


Assuntos
Mapeamento Cromossômico , Oryza , Raízes de Plantas , Locos de Características Quantitativas , Oryza/genética , Oryza/crescimento & desenvolvimento , Oryza/metabolismo , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Locos de Características Quantitativas/genética , Polimorfismo de Nucleotídeo Único/genética , Cromossomos de Plantas/genética , Genótipo
17.
Avian Pathol ; 42(4): 304-8, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23656603

RESUMO

Eimeria species parasites can cause the disease coccidiosis in all livestock species, most notably poultry. Traditional diagnostics such as faecal microscopy have now been supplemented by molecular assays including genus-specific and species-specific quantitative polymerase chain reaction (qPCR), although DNA extracted from faecal samples is commonly affected by PCR inhibition. This was confirmed when genomic DNA extracted from chicken faeces inhibited the threshold cycle value of internal positive control (IPC) DNA amplification by 15.33%. Hence, the objective of the present study was to use IPC qPCR to determine PCR inhibition in a series of experimental samples and use the increase in IPC qPCR threshold cycle value as an individual (sample-specific) correction factor for an established 5S rDNA qPCR used to estimate total Eimeria genome numbers. IPC-corrected genome counts were correlated with conventional oocyst per gram counts and compared with non-corrected counts, revealing a 0.1769 increase in correlation coefficient to outweigh underestimation of oocyst counts. Though the sample size used in this study is small, this limitation would be offset by the sample-specific correction factor determined using the IPC along with each sample.


Assuntos
Coccidiose/veterinária , Eimeria/genética , Genoma de Protozoário/genética , Doenças das Aves Domésticas/parasitologia , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Animais , Contagem de Células/veterinária , Fezes/química , Microscopia/veterinária , Oocistos , Reação em Cadeia da Polimerase/veterinária , RNA Ribossômico 5S/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Especificidade da Espécie
18.
Front Plant Sci ; 14: 1247014, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37731986

RESUMO

Biotic stress is one of the major threats to stable rice production. Climate change affects the shifting of pest outbreaks in time and space. Genetic improvement of biotic stress resistance in rice is a cost-effective and environment-friendly way to control diseases and pests compared to other methods such as chemical spraying. Fast deployment of the available and suitable genes/alleles in local elite varieties through marker-assisted selection (MAS) is crucial for stable high-yield rice production. In this review, we focused on consolidating all the available cloned genes/alleles conferring resistance against rice pathogens (virus, bacteria, and fungus) and insect pests, the corresponding donor materials, and the DNA markers linked to the identified genes. To date, 48 genes (independent loci) have been cloned for only major biotic stresses: seven genes for brown planthopper (BPH), 23 for blast, 13 for bacterial blight, and five for viruses. Physical locations of the 48 genes were graphically mapped on the 12 rice chromosomes so that breeders can easily find the locations of the target genes and distances among all the biotic stress resistance genes and any other target trait genes. For efficient use of the cloned genes, we collected all the publically available DNA markers (~500 markers) linked to the identified genes. In case of no available cloned genes yet for the other biotic stresses, we provided brief information such as donor germplasm, quantitative trait loci (QTLs), and the related papers. All the information described in this review can contribute to the fast genetic improvement of biotic stress resistance in rice for stable high-yield rice production.

19.
J Proteome Res ; 11(12): 5876-89, 2012 Dec 07.
Artigo em Inglês | MEDLINE | ID: mdl-23167294

RESUMO

Fasciolosis is an important foodborne, zoonotic disease of livestock and humans, with global annual health and economic losses estimated at several billion US$. Fasciola hepatica is the major species in temperate regions, while F. gigantica dominates in the tropics. In the absence of commercially available vaccines to control fasciolosis, increasing reports of resistance to current chemotherapeutic strategies and the spread of fasciolosis into new areas, new functional genomics approaches are being used to identify potential new drug targets and vaccine candidates. The glutathione transferase (GST) superfamily is both a candidate drug and vaccine target. This study reports the identification of a putatively novel Sigma class GST, present in a water-soluble cytosol extract from the tropical liver fluke F. gigantica. The GST was cloned and expressed as an enzymically active recombinant protein. This GST shares a greater identity with the human schistosomiasis GST vaccine currently at Phase II clinical trials than previously discovered F. gigantica GSTs, stimulating interest in its immuno-protective properties. In addition, in silico analysis of the GST superfamily of both F. gigantica and F. hepatica has revealed an additional Mu class GST, Omega class GSTs, and for the first time, a Zeta class member.


Assuntos
Fasciola/enzimologia , Glutationa Transferase/isolamento & purificação , Proteínas de Helminto/análise , Proteoma/análise , Proteômica/métodos , Sequência de Aminoácidos , Animais , Biologia Computacional/métodos , Citosol/enzimologia , Eletroforese em Gel Bidimensional , Ensaios Enzimáticos , Escherichia coli/genética , Fasciola/genética , Perfilação da Expressão Gênica , Glutationa Transferase/genética , Dados de Sequência Molecular , Filogenia , Análise Serial de Proteínas , Proteoma/genética , Proteínas Recombinantes/genética , Alinhamento de Sequência , Análise de Sequência de Proteína , Transformação Genética
20.
Nat Neurosci ; 11(3): 269-76, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18278043

RESUMO

Vertebrate cranial sensory ganglia, responsible for sensation of touch, taste and pain in the face and viscera, are composed of both ectodermal placode and neural crest cells. The cellular and molecular interactions allowing generation of complex ganglia remain unknown. Here, we show that proper formation of the trigeminal ganglion, the largest of the cranial ganglia, relies on reciprocal interactions between placode and neural crest cells in chick, as removal of either population resulted in severe defects. We demonstrate that ingressing placode cells express the Robo2 receptor and early migrating cranial neural crest cells express its cognate ligand Slit1. Perturbation of this receptor-ligand interaction by blocking Robo2 function or depleting either Robo2 or Slit1 using RNA interference disrupted proper ganglion formation. The resultant disorganization mimics the effects of neural crest ablation. Thus, our data reveal a novel and essential role for Robo2-Slit1 signaling in mediating neural crest-placode interactions during trigeminal gangliogenesis.


Assuntos
Movimento Celular/genética , Glicoproteínas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Crista Neural/embriologia , Receptores Imunológicos/metabolismo , Células-Tronco/metabolismo , Gânglio Trigeminal/embriologia , Animais , Comunicação Celular/genética , Diferenciação Celular/genética , Embrião de Galinha , Galinhas , Coturnix , Regulação para Baixo/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Glicoproteínas/genética , Proteínas do Tecido Nervoso/genética , Crista Neural/citologia , Crista Neural/metabolismo , Interferência de RNA , Receptores Imunológicos/genética , Células-Tronco/citologia , Gânglio Trigeminal/citologia , Gânglio Trigeminal/metabolismo , Proteínas Roundabout
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